Pesticidal genes and methods of use

ABSTRACT

Compositions having pesticidal activity and methods for their use are provided. Compositions include isolated and recombinant polypeptides having pesticidal activity, recombinant and synthetic nucleic acid molecules encoding the polypeptides, DNA constructs and vectors comprising the nucleic acid molecules, host cells comprising the vectors, and antibodies to the polypeptides. Nucleotide sequences encoding the polypeptides can be used in DNA constructs or expression cassettes for transformation and expression in organisms of interest. The compositions and methods provided are useful for producing organisms with enhanced pest resistance or tolerance. Transgenic plants and seeds comprising a nucleotide sequence that encodes a pesticidal protein of the invention are also provided. Such plants are resistant to insects and other pests. Methods are provided for producing the various polypeptides disclosed herein, and for using those polypeptides for controlling or killing a pest. Methods and kits for detecting polypeptides of the invention in a sample are also included.

CROSS REFERENCE TO RELATED APPLICATION

This application is a divisional application of U.S. application Ser. No. 16/184,769, filed Nov. 8, 2018, which is a divisional application of U.S. application Ser. No. 15/702,343, filed Sep. 12, 2017, now U.S. Pat. No. 10,167,324, issued Jan. 1, 2019, which is a divisional application of U.S. application Ser. No. 15/099,919, filed Apr. 15, 2016, now U.S. Pat. No. 10,053,707, issued Aug. 21, 2018, which claims the benefit of U.S. Provisional Application Ser. No. 62/151,156, filed Apr. 22, 2015, the contents of the applications are herein incorporated by reference in its entirety.

FIELD

The invention is drawn to methods and compositions for controlling pests, particularly plant pests.

REFERENCE TO A SEQUENCE LISTING SUBMITTED AS A TEXT FILE VIA EFS-WEB

The official copy of the sequence listing is submitted electronically via EFS-Web as an ASCII formatted sequence listing with a file named AgB012USDIV3_seqlisting.txt, created on Oct. 9, 2019, and having a size of 1.5 MB and is filed concurrently with the specification. The sequence listing contained in this ASCII formatted document is part of the specification and is herein incorporated by reference in its entirety.

BACKGROUND

Pests, plant diseases, and weeds can be serious threats to crops. Losses due to pests and diseases have been estimated at 37% of the agricultural production worldwide, with 13% due to insects, bacteria and other organisms.

Toxins are virulence determinants that play an important role in microbial pathogenicity and/or evasion of the host immune response. Toxins from the gram-positive bacterium Bacillus, particularly Bacillus thuringiensis, have been used as insecticidal proteins. Current strategies use the genes expressing these toxins to produce transgenic crops. Transgenic crops expressing insecticidal protein toxins are used to combat crop damage from insects.

While the use of Bacillus toxins has been successful in controlling insects, resistance to Bt toxins has developed in some target pests in many parts of the world where such toxins have been used intensively. One way of solving this problem is sowing Bt crops with alternating rows of regular non Bt crops (refuge). An alternative method to avoid or slow down development of insect resistance is stacking insecticidal genes with different modes of action against insects in transgenic plants. The current strategy of using transgenic crops expressing insecticidal protein toxins is placing increasing emphasis on the discovery of novel toxins, beyond those already derived from the bacterium Bacillus thuringiensis. These toxins may prove useful as alternatives to those derived from B. thuringiensis for deployment in insect- and pest-resistant transgenic plants. Thus, new toxin proteins are needed.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 provides an alignment of SEQ ID NOS: 177, 307, 372, 374, and 396. Divergent amino acids are highlighted in grey.

FIG. 2 provides the global sequence identity relationship between SEQ ID NOS: 177, 307, 372, 374, and 396.

SUMMARY

Compositions having pesticidal activity and methods for their use are provided. Compositions include isolated and recombinant polypeptide sequences having pesticidal activity, recombinant and synthetic nucleic acid molecules encoding the pesticidal polypeptides, DNA constructs comprising the nucleic acid molecules, vectors comprising the nucleic acid molecules, host cells comprising the vectors, and antibodies to the pesticidal polypeptides. Nucleotide sequences encoding the polypeptides provided herein can be used in DNA constructs or expression cassettes for transformation and expression in organisms of interest, including microorganisms and plants.

The compositions and methods provided herein are useful for the production of organisms with enhanced pest resistance or tolerance. These organisms and compositions comprising the organisms are desirable for agricultural purposes. Transgenic plants and seeds comprising a nucleotide sequence that encodes a pesticidal protein of the invention are also provided. Such plants are resistant to insects and other pests.

Methods are provided for producing the various polypeptides disclosed herein, and for using those polypeptides for controlling or killing a pest. Methods and kits for detecting polypeptides of the invention in a sample are also included.

DETAILED DESCRIPTION

The present inventions now will be described more fully hereinafter with reference to the accompanying drawings, in which some, but not all embodiments of the inventions are shown. Indeed, these inventions may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. Like numbers refer to like elements throughout.

Many modifications and other embodiments of the inventions set forth herein will come to mind to one skilled in the art to which these inventions pertain having the benefit of the teachings presented in the foregoing descriptions and the associated drawings. Therefore, it is to be understood that the inventions are not to be limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims. Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

I. Polynucleotides and Polypeptides

Compositions and method for conferring pesticidal activity to an organism are provided. The modified organism exhibits pesticidal resistance or tolerance.

Recombinant pesticidal proteins, or polypeptides and fragments and variants thereof that retain pesticidal activity, are provided and include those set forth in SEQ ID NOs: 1-398. The pesticidal proteins are biologically active (e.g., pesticidal) against pests including insects, fungi, nematodes, and the like. Nucleotides encoding the pesticidal polypeptides, including for example, SEQ ID NOS: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398 or active fragments or variants thereof, can be used to produce transgenic organisms, such as plants and microorganisms. The pesticidal proteins are biologically active (for example, are pesticidal) against pests including insects, fungi, nematodes, and the like. Polynucleotides encoding the pesticidal polypeptides, including for example, SEQ ID NOS: 1-398 or active fragments or variants thereof, can be used to produce transgenic organisms, such as plants and microorganisms. The transformed organisms are characterized by genomes that comprise at least one stably incorporated DNA construct comprising a coding sequence for a pesticidal protein disclosed herein. In some embodiments, the coding sequence is operably linked to a promoter that drives expression of the encoded pesticidal polypeptide. Accordingly, transformed microorganisms, plant cells, plant tissues, plants, seeds, and plant parts are provided. A summary of various polypeptides, active variants and fragments thereof, and polynucleotides encoding the same are set forth below in Table 1. As noted in Table 1, various forms of polypeptides are provided. Full length pesticidal polypeptides, as well as, modified versions of the original full-length sequence (i.e., variants) are provided. Table 1 further denotes “CryBP1” sequences. Such sequences comprise accessory polypeptides that can be associated with some of the toxin genes. In such instances, the CryBP1 sequences can be used alone or in combination with any of the pesticidal polypeptides provided herein. Table 1 further provides Split-Cry C-terminus polypeptides. Such sequences comprise the sequence of a downstream protein that has homology to the C-terminal end of the Cry class of toxin genes and are usually found after a Cry gene that is not full-length and is missing the expected C-terminal region.

TABLE 1 Summary of SEQ ID NOs, Gene Class, and Variants thereof Polypeptides of the invention Polypeptides of the invention Split-Cry (and polynucleotides (and polynucleotides Full- Modified CryBP1 C-terminus encoding the same) include encoding the same) include Gene length SEQID SEQ ID SEQ ID Gene those having the % sequence those having the similarity set Name SEQ ID No. No.(s) No. No. Homologs Class identity listed below forth below APG00056 1 2 AGU13855.1 (69.6% identity, 79.2% similarity) Cry32 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 AGU13869.1 (68.4% identity, 79.1% similarity) US20110203014_23 (68.4% identity, 79.1% similarity) APG00058 (63.6% identity, 75.5% similarity) Cry32Ea1 (55.1% identity, 65.9% similarity) APG00068 (52.0% identity, 64.2% similarity) APG00105 (50.5% identity, 61.8% similarity) APG00054 (50.3% identity, 60.9% similarity) APG00058 3 4 AGU13869.1 (74.6% identity, 81.5% similarity) Cry32 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 US20110203014_23 (74.6% identity, 81.5% similarity) AGU13855.1 (69.3% identity, 77.8% similarity) APG00056 (63.6% identity, 75.5% similarity) Cry32Da1 (52.7% identity, 64.1% similarity) APG00064 5 6 US20130227743A1_58 (66.4% identity, 78.7% similarity) Mtx 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_000875422.1 (65.8% identity, 79.6% similarity) US20060191034A1_10 (65.8% identity, 78.7% similarity) WP_000875423.1 (65.2% identity, 79.3% similarity) APG00055 (64.2% identity, 78.5% similarity) APG00174 (61.6% identity, 74.9% similarity) APG00768 (57.9% identity, 73.5% similarity) APG00067 7 APG00408 (81.6% identity, 89.3% similarity) Mtx 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00559 (75.5% identity, 81.2% similarity) APG00006 (74.4% identity, 85.9% similarity) APG00201 (74.4% identity, 84.9% similarity) WP_000963933.1 (72.6% identity, 84.3% similarity) US20130227743A1_100 (72.5% identity, 83.2% similarity) APG00155 (71.8% identity, 81.5% similarity) APG00107 (71.5% identity, 83.6% similarity) APG00022 (71.1% identity, 83.7% similarity) APG00036 (70.9% identity, 80.5% similarity) US20130227743A1_60 (40.7% identity, 46.9% similarity) WP_026324166.1 (26.9% identity, 42.7% similarity) APG00071 8 9 APG00150 (95.9% identity, 98.6% similarity) Vip 25, 30, 35, 40, 45, 50, 55, 60, 35, 40, 45, 50, 55, 60, 65, 70, 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 CAI96522.1 (22.2% identity, 34.9% similarity) US_6369213_B1-94 (22.2% identity, 34.9% similarity) AFJ19238.1 (22.2% identity, 34.8% similarity) Vip3Aa49 (22.2% identity, 34.7% similarity) APG00073 10 11, 12, 13 CR5AC_BACTU (64.9% identity, 71.0% similarity) Cry5 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 US_5281530-3 (64.4% identity, 71.9% similarity) Cry5Ab1 (64.4% identity, 71.8% similarity) APG00074 14 228 KEZ80024.1 (39.4% identity, 48.8% similarity) Vip 40, 45, 50, 55, 60, 65, 70, 75, 50, 55, 60, 65, 70, 75, 80, 85, 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 Vip3Ag2 (23.6% identity, 40.2% similarity) AIT93180.1 (23.6% identity, 40.1% similarity) APG00084 15  16, 229 APG00152 (59.4% identity, 69.8% similarity) Cry 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 US20130227743A1_12 (57.6% identity, 70.5% similarity) WP_003308604.1 (55.4% identity, 66.8% similarity) APG00164 (53.6% identity, 68.1% similarity) CBL59394.1 (46.2% identity, 63.8% similarity) Cry8Aa1 (28.8% identity, 38.0% similarity) APG00105 17 18, 19, 20 AGU13822.1 (78.4% identity, 84.5% similarity) Cry32 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 AGU13821.1 (57.0% identity, 69.8% similarity) AGU13871.1 (56.1% identity, 68.2% similarity) Cry32Ab1 (54.2% identity, 66.7% similarity) APG00054 (53.4% identity, 65.2% similarity) APG00068 (52.1% identity, 65.4% similarity) APG00056 (50.5% identity, 61.8% similarity) APG00107 21 APG00155 (78.0% identity, 87.5% similarity) Mtx 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_000963933.1 (77.4% identity, 88.9% similarity) US20130227743A1_100 (77.0% identity, 87.8% similarity) APG00201 (75.3% identity, 83.6% similarity) APG00006 (74.9% identity, 84.4% similarity) APG00067 (71.5% identity, 83.6% similarity) APG00036 (71.3% identity, 80.0% similarity) APG00022 (69.8% identity, 80.4% similarity) APG00137 (69.4% identity, 81.2% similarity) APG00898 (64.8% identity, 78.8% similarity) US20130227743A1_60 (39.0% identity, 44.9% similarity) WP_012181803.1 (23.0% identity, 37.8% similarity) APG00108 22 230 WP_030313032.1 (71.8% identity, 72.2% similarity) Cyt 75, 80, 85, 90, 95, 96, 97, 98, 75, 80, 85, 90, 95, 96, 97, 98, 99 99 WP_015038174.1 (69.5% identity, 71.1% similarity) APG00134 (68.8% identity, 69.9% similarity) WP_030212110.1 (66.9% identity, 69.5% similarity) APG00198 (62.0% identity, 66.5% similarity) Cyt2Ca1 (19.9% identity, 34.7% similarity) APG00112 23 AGA40062.1 (80.6% identity, 88.3% similarity) Mtx 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 AGA40061.1 (61.3% identity, 73.9% similarity) APG00907 (57.7% identity, 69.5% similarity) Cry60Ba1 (36.2% identity, 51.6% similarity) APG00113 24 231, 232 APG00196 (91.0% identity, 95.3% similarity) Cry 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00030 (71.4% identity, 84.0% similarity) AGP17989.1 (68.7% identity, 82.6% similarity) APG00096 (68.1% identity, 82.2% similarity) APG00114 (58.5% identity, 77.2% similarity) AGP17990.1 (57.9% identity, 68.4% similarity) APG00500 (56.6% identity, 65.3% similarity) US_2013_0227743_A1_194 (41.3% identity, 47.4% similarity) Cry42Aa1 (23.6% identity, 37.1% similarity) APG00116 25  26, 233, 234 AGP18043.1 (94.2% identity, 96.6% similarity) Cry 95, 96, 97, 98, 99 97, 98, 99 AFM37573.1 (93.9% identity, 96.1% similarity) AGV55020.1 (86.8% identity, 92.6% similarity) APG00153 (53.8% identity, 64.4% similarity) Cry53Aa1 (39.8% identity, 54.8% similarity) APG00117 27 235, 236 APG00186 (97.2% identity, 97.4% similarity) Cyt 35, 40, 45, 50, 55, 60, 65, 70, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 96, 97, 98, 75, 80, 85, 90, 95, 96, 97, 98, 99 99 AJF68767.1 (31.8% identity, 32.3% similarity) WP_030689537.1 (31.5% identity, 32.2% similarity) WP_030313032.1 (30.4% identity, 32.0% similarity) Cyt2Aa2 (10.5% identity, 18.6% similarity) APG00118 28 237 APG00223 (91.0% identity, 95.2% similarity) Bin 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00913 (90.9% identity, 92.7% similarity) APG00454 (90.3% identity, 94.9% similarity) APG00242 (90.3% identity, 94.7% similarity) WP_002187944.1 (84.6% identity, 86.9% similarity) US20130227743A1_10 (81.8% identity, 86.2% similarity) WP_001258160.1 (81.6% identity, 85.9% similarity) APG00065 (58.1% identity, 70.6% similarity) Cry35Ba1 (18.8% identity, 32.5% similarity) APG00121 29 238 APG00177 (68.6% identity, 79.9% similarity) Cyt 55, 60, 65, 70, 75, 80, 85, 90, 70, 75, 80, 85, 90, 95, 96, 97, 95, 96, 97, 98, 99 98, 99 APG00126 (54.7% identity, 68.1% similarity) WP_016110459.1 (54.3% identity, 63.7% similarity) APG00128 (54.3% identity, 63.7% similarity) WP_016110460.1 (51.8% identity, 66.4% similarity) CT2BB_BACTJ (38.9% identity, 52.5% similarity) Cyt2Ca1 (37.6% identity, 50.9% similarity) APG00131 30 Vip3Ag5 (29.9% identity, 44.1% similarity) Vip 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00132 31 239 APG00249 (88.9% identity, 94.7% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00459 (88.0% identity, 94.1% similarity) APG00065 (77.7% identity, 85.0% similarity) APG00407 (75.9% identity, 85.2% similarity) APG00229 (73.7% identity, 82.4% similarity) WP_000839920.1 (71.9% identity, 82.9% similarity) WP_002166959.1 (68.4% identity, 79.7% similarity) US20130227743A1_146 (68.2% identity, 79.4% similarity) Cry35Ab3 (20.7% identity, 35.2% similarity) APG00134 32 240 WP_030313032.1 (94.8% identity, 96.9% similarity) Cyt 95, 96, 97, 98, 99 97, 98, 99 WP_015038174.1 (93.8% identity, 96.4% similarity) WP_030212110.1 (93.2% identity, 96.4% similarity) APG00198 (86.5% identity, 92.2% similarity) APG00108 (68.8% identity, 69.9% similarity) Cyt2Ca1 (24.2% identity, 42.0% similarity) APG00137 33 241 APG00006 (71.8% identity, 83.6% similarity) Mtx 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00201 (71.7% identity, 81.2% similarity) WP_000963933.1 (71.6% identity, 82.5% similarity) US20130227743A1_100 (70.0% identity, 81.5% similarity) APG00107 (69.4% identity, 81.2% similarity) APG00898 (69.2% identity, 80.0% similarity) APG00067 (67.2% identity, 81.6% similarity) APG00036 (67.2% identity, 78.0% similarity) APG00408 (66.2% identity, 78.0% similarity) APG00022 (65.2% identity, 77.0% similarity) US20130227743A1_60 (39.2% identity, 46.7% similarity) WP_012181803.1 (24.5% identity, 36.9% similarity) APG00141 34 242 WP_000727408.1 (93.5% identity, 96.6% similarity) Bin 95, 96, 97, 98, 99 97, 98, 99 WP_016110923.1 (93.2% identity, 96.3% similarity) WP_000727409.1 (92.7% identity, 95.8% similarity) APG00237 (61.2% identity, 71.8% similarity) APG00261 (60.5% identity, 71.5% similarity) APG00913 (54.0% identity, 65.1% similarity) APG00065 (53.4% identity, 67.1% similarity) APG00398 (50.8% identity, 60.9% similarity) Cry35Ab5 (22.2% identity, 38.4% similarity) APG00150 35 243 APG00071 (95.9% identity, 98.6% similarity) Vip 25, 30, 35, 40, 45, 50, 55, 60, 35, 40, 45, 50, 55, 60, 65, 70, 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 Vip3Aa49 (22.4% identity, 34.8% similarity) APG00152 36 37 US20130227743A1_12 (69.7% identity, 80.8% similarity) Cry 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_003308604.1 (66.0% identity, 75.9% similarity) APG00084 (59.4% identity, 69.8% similarity) D5H3I8_BACTG (45.3% identity, 62.9% similarity) Cry8Aa1 (27.8% identity, 37.8% similarity) APG00153 38  39, 244, 245 AGP18043.1 (54.1% identity, 65.0% similarity) Cry 55, 60, 65, 70, 75, 80, 85, 90, 70, 75, 80, 85, 90, 95, 96, 97, 95, 96, 97, 98, 99 98, 99 AFM37573.1 (53.9% identity, 64.7% similarity) APG00116 (53.8% identity, 64.4% similarity) AGV55020.1 (53.4% identity, 65.3% similarity) Cry8Ba1 (21.6% identity, 33.6% similarity) APG00155 40 WP_000963933.1 (78.5% identity, 85.9% similarity) Mtx 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00107 (78.0% identity, 87.5% similarity) US20130227743A1_100 (77.0% identity, 85.1% similarity) APG00201 (76.3% identity, 83.9% similarity) APG00006 (75.7% identity, 83.0% similarity) APG00036 (74.8% identity, 82.4% similarity) APG00022 (72.1% identity, 80.7% similarity) APG00067 (71.8% identity, 81.5% similarity) APG00898 (67.1% identity, 79.3% similarity) APG00137 (66.4% identity, 77.6% similarity) US20130227743A1_60 (41.6% identity, 46.9% similarity) WP_000794513.1 (22.9% identity, 35.3% similarity) APG00164 41 42 CBL59393.1 (58.2% identity, 73.3% similarity) Cry 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 D5H3J0_BACTG (58.2% identity, 73.3% similarity) D5H3I8_BACTG (58.0% identity, 73.2% similarity) APG00084 (53.6% identity, 68.1% similarity) Cry8Aa1 (37.2% identity, 43.6% similarity) APG00166 43 246 KEZ80024.1 (34.1% identity, 37.7% similarity) Vip 35, 40, 45, 50, 55, 60, 65, 70, 40, 45, 50, 55, 60, 65, 70, 75, 75, 80, 85, 90, 95, 96, 97, 98, 80, 85, 90, 95, 96, 97, 98, 99 99 US_7129212_B2-19 (17.5% identity, 28.4% similarity) US_7129212_B2-17 (17.1% identity, 28.4% similarity) Vip3Ag2 (16.8% identity, 27.9% similarity) APG00168 44 247 APG00015 (60.3% identity, 71.4% similarity) Cyt2 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 Cyt2Aa2 (59.5% identity, 71.8% similarity) APG00172 45 AHN21707.1 (72.8% identity, 84.3% similarity) Mtx 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_024361905.1 (72.8% identity, 84.3% similarity) WP_020721402.1 (58.1% identity, 74.8% similarity) Cry46Ab (37.2% identity, 52.6% similarity) APG00173 46 47 APG00278 (94.5% identity, 96.8% similarity) Vip 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00077 (82.4% identity, 87.7% similarity) APG00175 (80.5% identity, 86.0% similarity) KEZ80024.1 (73.6% identity, 84.3% similarity) APG00273 (68.8% identity, 78.1% similarity) APG00104 (68.7% identity, 78.1% similarity) APG00033 (68.1% identity, 77.4% similarity) Vip3Aa13 (24.3% identity, 41.6% similarity) APG00175 48 49 APG00077 (97.4% identity, 97.9% similarity) Vip 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00278 (80.7% identity, 86.6% similarity) APG00173 (80.5% identity, 86.0% similarity) APG00273 (78.0% identity, 85.7% similarity) APG00033 (76.9% identity, 84.7% similarity) APG00104 (71.3% identity, 80.3% similarity) KEZ80024.1 (68.1% identity, 78.7% similarity) Vip3Ai1 (25.0% identity, 42.7% similarity) APG00176 50 51 AGP18009.1 (44.3% identity, 58.8% similarity) Cry 45, 50, 55, 60, 65, 70, 75, 80, 60, 65, 70, 75, 80, 85, 90, 95, 85, 90, 95, 96, 97, 98, 99 96, 97, 98, 99 AGP17994.1 (44.2% identity, 58.3% similarity) US_8461415_B2-49 (42.2% identity, 56.1% similarity) Cry53Aa1 (34.7% identity, 49.1% similarity) APG00177 52  53, 248 APG00121 (68.6% identity, 79.9% similarity) Cyt 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 WP_016110459.1 (56.8% identity, 68.9% similarity) APG00128 (56.6% identity, 69.9% similarity) APG00126 (56.5% identity, 73.1% similarity) WP_016110460.1 (55.4% identity, 72.5% similarity) US_6686452-2 (42.1% identity, 54.5% similarity) Cyt2Ca1 (41.7% identity, 54.1% similarity) APG00178 54 APG00727 (90.7% identity, 93.6% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00315 (87.7% identity, 91.6% similarity) Cry49Ab1 (27.7% identity, 44.2% similarity) APG00180 55 WP_000240776.1 (79.3% identity, 91.2% similarity) Cry6 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_000240775.1 (79.0% identity, 90.7% similarity) WP_016078427.1 (79.0% identity, 90.4% similarity) APG00024 (77.7% identity, 88.9% similarity) APG00342 (62.9% identity, 78.8% similarity) Cry6Ba1 (27.2% identity, 48.1% similarity) APG00186 56 249, 250 APG00117 (97.2% identity, 97.4% similarity) Cyt 35, 40, 45, 50, 55, 60, 65, 70, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 96, 97, 98, 75, 80, 85, 90, 95, 96, 97, 98, 99 99 AJF68767.1 (31.8% identity, 32.3% similarity) WP_030689537.1 (31.5% identity, 32.2% similarity) WP_030313032.1 (30.4% identity, 32.0% similarity) Cyt2Aa2 (10.6% identity, 18.8% similarity) APG00188 57 58 APG00007 (76.5% identity, 83.7% similarity) Cry 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 AGP18022.1 (74.9% identity, 83.0% similarity) AEH76820.1 (44.4% identity, 59.2% similarity) US20130227743A1_48 (44.3% identity, 58.5% similarity) Cry32Ca1 (44.3% identity, 57.3% similarity) APG00189 59 60, 61 APG00087 (90.3% identity, 94.0% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US_8759619_B2-30 (28.0% identity, 43.2% similarity) US_7253343_B2-17 (27.7% identity, 41.1% similarity) Cry1Ib4 (27.3% identity, 40.9% similarity) APG00190 62 63 WP_001083588.1 (69.3% identity, 79.0% similarity) Cry26 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 AGT50919.1 (68.1% identity, 78.0% similarity) Cry26Aa1 (65.4% identity, 75.3% similarity) APG00192 64 WP_002167240.1 (32.2% identity, 51.3% similarity) Bin 35, 40, 45, 50, 55, 60, 65, 70, 55, 60, 65, 70, 75, 80, 85, 90, 75, 80, 85, 90, 95, 96, 97, 98, 95, 96, 97, 98, 99 99 WP_002016877.1 (32.2% identity, 49.6% similarity) WP_016097060.1 (31.3% identity, 49.0% similarity) Cry35Ab2 (23.3% identity, 38.3% similarity) APG00194 65 251 APG00017 (57.4% identity, 72.2% similarity) Cyt 40, 45, 50, 55, 60, 65, 70, 75, 55, 60, 65, 70, 75, 80, 85, 90, 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 US_8513493_B2-47 (35.7% identity, 53.1% similarity) 2RCI_A (27.0% identity, 43.8% similarity) CT2BB_BACTJ (27.0% identity, 40.9% similarity) Cyt2Aa2 (26.9% identity, 42.8% similarity) APG00196 66 252 APG00113 (91.0% identity, 95.3% similarity) Cry 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00030 (69.9% identity, 82.5% similarity) AGP17989.1 (66.9% identity, 81.1% similarity) APG00096 (66.6% identity, 80.9% similarity) APG00114 (57.1% identity, 76.0% similarity) AGP17990.1 (56.6% identity, 67.0% similarity) APG00500 (51.7% identity, 62.1% similarity) US_2013_0227743_A1_194 (39.6% identity, 46.0% similarity) Cry42Aa1 (23.4% identity, 36.7% similarity) APG00198 67 253 WP_024758398.1 (91.8% identity, 95.9% similarity) Cyt 95, 96, 97, 98, 99 96, 97, 98, 99 WP_030208963.1 (90.7% identity, 94.3% similarity) WP_030545473.1 (89.8% identity, 92.9% similarity) APG00134 (86.5% identity, 92.2% similarity) APG00108 (62.0% identity, 66.5% similarity) Cyt2Ca1 (22.9% identity, 40.3% similarity) APG00200 68 254, 255 APG00186 (53.9% identity, 58.7% similarity) Cyt 40, 45, 50, 55, 60, 65, 70, 75, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 96, 97, 98, 99 80, 85, 90, 95, 96, 97, 98, 99 APG00117 (52.6% identity, 57.5% similarity) WP_030752998.1 (35.8% identity, 37.7% similarity) WP_030497750.1 (35.2% identity, 37.1% similarity) WP_031015242.1 (35.0% identity, 37.3% similarity) Cyt2Aa2 (11.7% identity, 21.5% similarity) APG00203 69 256 APG00353 (95.3% identity, 96.6% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00301 (90.9% identity, 93.7% similarity) APG00844 (89.8% identity, 93.7% similarity) APG00412 (87.5% identity, 91.3% similarity) APG00065 (75.7% identity, 83.2% similarity) WP_000839920.1 (73.0% identity, 82.8% similarity) WP_002166959.1 (70.4% identity, 80.9% similarity) WP_002191947.1 (70.2% identity, 80.9% similarity) Cry35Ad2 (22.9% identity, 36.8% similarity) APG00207 70 Vip3Aa8 (37.8% identity, 53.6% similarity) Vip 40, 45, 50, 55, 60, 65, 70, 75, 55, 60, 65, 70, 75, 80, 85, 90, 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00209 71 257 WP_002128069.1 (73.1% identity, 85.5% similarity) Cry6 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 WP_000591974.1 (73.1% identity, 85.2% similarity) WP_002185458.1 (73.1% identity, 85.2% similarity) WP_002191530.1 (73.1% identity, 85.2% similarity) APG00210 72 258 WP_028939439.1 (59.6% identity, 74.1% similarity) Cry6 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 WP_015096649.1 (42.6% identity, 63.0% similarity) WP_019823923.1 (32.2% identity, 51.3% similarity) WP_016779464.1 (32.0% identity, 51.8% similarity) APG00211 73 74 APG00368 (98.7% identity, 99.2% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00011 (96.2% identity, 97.3% similarity) APG00377 (79.1% identity, 87.4% similarity) APG00356 (79.1% identity, 86.9% similarity) APG00231 (78.6% identity, 86.9% similarity) APG00035 (76.9% identity, 85.5% similarity) WP_000143307.1 (76.4% identity, 85.8% similarity) WP_000143308.1 (75.1% identity, 85.3% similarity) US20130227743A1_6 (69.4% identity, 79.4% similarity) Cry35Ab4 (22.7% identity, 38.5% similarity) APG00212 75 259, 260 APG00798 (92.7% identity, 96.8% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 APG00592 (89.8% identity, 94.2% similarity) APG00600 (87.1% identity, 92.0% similarity) APG00619 (86.2% identity, 92.0% similarity) US20130227743A1_146 (76.9% identity, 83.3% similarity) APG00065 (72.0% identity, 80.7% similarity) WP_002191947.1 (71.2% identity, 79.2% similarity) WP_002166959.1 (70.9% identity, 79.2% similarity) Cry49Aa1 (22.4% identity, 34.4% similarity) APG00213 76 261 APG00243 (82.7% identity, 89.1% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00844 (81.4% identity, 89.9% similarity) APG00203 (81.2% identity, 90.2% similarity) APG00353 (80.4% identity, 89.4% similarity) APG00065 (79.2% identity, 84.1% similarity) WP_000839920.1 (72.3% identity, 83.0% similarity) WP_002166959.1 (71.1% identity, 81.2% similarity) WP_002191947.1 (70.8% identity, 81.2% similarity) Cry35Ad2 (21.5% identity, 37.1% similarity) APG00214 77 262 WP_033669474.1 (90.4% identity, 95.6% similarity) Cry6 95, 96, 97, 98, 99 96, 97, 98, 99 EOP91866.1 (84.6% identity, 89.6% similarity) KFN04132.1 (73.4% identity, 86.0% similarity) WP_016092834.1 (72.8% identity, 84.6% similarity) APG00215 78 WP_016093722.1 (73.0% identity, 82.5% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_002167240.1 (59.6% identity, 72.3% similarity) APG00386 (59.0% identity, 73.7% similarity) WP_002090518.1 (53.4% identity, 70.7% similarity) Cry35Ad2 (22.8% identity, 40.5% similarity) APG00216 79 APG00090 (73.0% identity, 73.3% similarity) Bin 55, 60, 65, 70, 75, 80, 85, 90, 70, 75, 80, 85, 90, 95, 96, 97, 95, 96, 97, 98, 99 98, 99 US20130227743A1_154 (54.1% identity, 69.7% similarity) US20130227743A1_156 (54.1% identity, 66.3% similarity) KEZ80012.1 (44.3% identity, 55.4% similarity) Cry49Ab1 (17.6% identity, 27.5% similarity) APG00219 80 263 AGA40057.1 (33.9% identity, 51.3% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 55, 60, 65, 70, 75, 80, 85, 90, 75, 80, 85, 90, 95, 96, 97, 98, 95, 96, 97, 98, 99 99 AGA40058.1 (29.9% identity, 46.9% similarity) WP_017762619.1 (29.7% identity, 43.3% similarity) WP_017762581.1 (29.5% identity, 44.7% similarity) APG00221 81 264 APG00376 (92.5% identity, 95.1% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00237 (88.1% identity, 91.6% similarity) APG00353 (77.0% identity, 84.8% similarity) APG00844 (75.7% identity, 83.7% similarity) WP_002166959.1 (67.7% identity, 79.1% similarity) WP_000839920.1 (67.4% identity, 79.5% similarity) WP_002191947.1 (67.4% identity, 79.1% similarity) APG00065 (63.5% identity, 75.0% similarity) Cry35Ac2 (21.9% identity, 39.4% similarity) APG00223 82 265, 266 APG00242 (98.4% identity, 98.6% similarity) Bin 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00454 (97.7% identity, 98.4% similarity) APG00118 (91.0% identity, 95.2% similarity) US20130227743A1_10 (85.6% identity, 88.7% similarity) WP_001258160.1 (85.4% identity, 88.4% similarity) WP_001258161.1 (85.2% identity, 88.2% similarity) APG00913 (85.1% identity, 90.0% similarity) APG00065 (58.3% identity, 70.5% similarity) Cry35Ab4 (19.1% identity, 34.1% similarity) APG00227 83 267, 268 APG00517 (79.8% identity, 79.8% similarity) Mtx 55, 60, 65, 70, 75, 80, 85, 90, 65, 70, 75, 80, 85, 90, 95, 96, 95, 96, 97, 98, 99 97, 98, 99 APG00635 (55.0% identity, 66.2% similarity) APG00020 (53.3% identity, 64.4% similarity) WP_002166885.1 (52.2% identity, 61.8% similarity) US20130227743A1_110 (39.4% identity, 54.0% similarity) AGP17985.1 (34.3% identity, 49.0% similarity) US_8829279_B2-11 (30.9% identity, 44.1% similarity) APG00229 84 269, 270 APG00419 (92.6% identity, 94.1% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00065 (86.9% identity, 92.8% similarity) APG00844 (86.4% identity, 89.1% similarity) APG00301 (85.4% identity, 88.4% similarity) APG00412 (84.9% identity, 89.1% similarity) WP_000839920.1 (68.1% identity, 77.5% similarity) WP_002166959.1 (67.6% identity, 77.6% similarity) WP_002191947.1 (67.3% identity, 77.6% similarity) Cry35Ab3 (21.9% identity, 34.9% similarity) APG00230 85 271 APG00413 (81.0% identity, 88.9% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 KEZ80012.1 (79.9% identity, 87.6% similarity) WP_017154552.1 (72.3% identity, 83.0% similarity) APG00596 (69.1% identity, 80.9% similarity) APG00191 (67.5% identity, 81.0% similarity) APG00741 (65.8% identity, 78.2% similarity) APG00757 (62.3% identity, 73.8% similarity) APG00090 (59.9% identity, 77.2% similarity) US20130227743A1_156 (51.3% identity, 58.4% similarity) Cry35Ad2 (21.3% identity, 36.7% similarity) APG00231 86 272 APG00377 (96.8% identity, 98.7% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00356 (95.4% identity, 98.1% similarity) APG00287 (94.4% identity, 97.3% similarity) APG00731 (89.3% identity, 93.6% similarity) APG00035 (87.7% identity, 93.3% similarity) WP_000143307.1 (87.7% identity, 91.7% similarity) WP_000143308.1 (84.5% identity, 91.2% similarity) US20130227743A1_6 (79.4% identity, 83.9% similarity) APG00011 (77.7% identity, 85.5% similarity) Cry35Ab4 (22.4% identity, 39.0% similarity) APG00232 87 273 AGA40057.1 (27.5% identity, 41.5% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US20130227743A1_200 (26.7% identity, 41.2% similarity) AGP17992.1 (24.4% identity, 35.9% similarity) Cry73Aa (20.0% identity, 31.5% similarity) APG00233 88 274 APG00251 (81.8% identity, 87.8% similarity) Cry 40, 45, 50, 55, 60, 65, 70, 75, 55, 60, 65, 70, 75, 80, 85, 90, 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 US20130227743A1_200 (38.7% identity, 52.6% similarity) AGA40058.1 (25.4% identity, 38.9% similarity) WP_017762619.1 (25.0% identity, 40.7% similarity) Cry41Ba2 (13.1% identity, 20.4% similarity) APG00235 89 275 US20130227743A1_170 (32.8% identity, 37.8% CRY 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, similarity) 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 US20130227743A1_200 (29.8% identity, 41.7% similarity) WP_017762619.1 (29.2% identity, 45.7% similarity) Cyt1Da1 (17.9% identity, 33.1% similarity) APG00237 90 276 APG00221 (88.1% identity, 91.6% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00376 (85.3% identity, 89.8% similarity) APG00203 (77.5% identity, 82.8% similarity) APG00844 (77.3% identity, 82.6% similarity) WP_002166959.1 (69.1% identity, 78.9% similarity) WP_002191947.1 (68.8% identity, 78.9% similarity) WP_000839920.1 (68.6% identity, 79.6% similarity) APG00065 (65.0% identity, 75.1% similarity) Cry35Ab3 (22.7% identity, 38.7% similarity) APG00239 91 277 WP_017762619.1 (27.3% identity, 38.4% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_017762581.1 (26.9% identity, 37.3% similarity) AGA40058.1 (26.8% identity, 41.8% similarity) AGP17992.1 (26.2% identity, 38.6% similarity) APG00241 92 278, 279 KFN03392.1 (55.9% identity, 71.4% similarity) Cry6 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 WP_033674722.1 (55.2% identity, 72.8% similarity) WP_003196831.1 (54.5% identity, 71.2% similarity) WP_018765384.1 (54.5% identity, 71.2% similarity) APG00298 (52.0% identity, 71.0% similarity) APG00242 93 280 APG00223 (98.4% identity, 98.6% similarity) Bin 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00454 (97.9% identity, 98.8% similarity) APG00118 (90.3% identity, 94.7% similarity) US20130227743A1_10 (85.2% identity, 88.4% similarity) WP_001258160.1 (85.0% identity, 88.2% similarity) WP_001258161.1 (84.7% identity, 88.0% similarity) APG00913 (84.5% identity, 89.6% similarity) APG00065 (58.8% identity, 71.4% similarity) Cry35Ab3 (19.3% identity, 34.5% similarity) APG00243 94 281 APG00301 (86.7% identity, 92.7% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 APG00213 (82.7% identity, 89.1% similarity) APG00844 (82.0% identity, 90.1% similarity) APG00412 (81.6% identity, 89.3% similarity) APG00065 (80.4% identity, 85.1% similarity) WP_000839920.1 (77.4% identity, 84.3% similarity) WP_002166959.1 (73.1% identity, 81.4% similarity) WP_002191947.1 (72.9% identity, 81.4% similarity) Cry35Ad2 (23.0% identity, 37.5% similarity) APG00248 95 282 APG00262 (63.2% identity, 73.2% similarity) Bin 60, 65, 70, 75, 80, 85, 90, 95, 70, 75, 80, 85, 90, 95, 96, 97, 96, 97, 98, 99 98, 99 WP_002191947.1 (59.5% identity, 69.9% similarity) WP_002166959.1 (59.3% identity, 69.9% similarity) APG00243 (59.1% identity, 69.9% similarity) AGA40039.1 (58.0% identity, 69.2% similarity) APG00369 (57.6% identity, 68.1% similarity) APG00212 (57.1% identity, 67.6% similarity) APG00065 (56.8% identity, 69.6% similarity) Cry35Ab1 (20.6% identity, 33.7% similarity) APG00249 96 283, 284 APG00459 (89.0% identity, 92.8% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00132 (88.9% identity, 94.7% similarity) APG00065 (77.4% identity, 83.8% similarity) APG00407 (75.5% identity, 84.4% similarity) APG00229 (73.0% identity, 81.3% similarity) WP_000839920.1 (71.7% identity, 81.0% similarity) WP_002166959.1 (68.3% identity, 78.0% similarity) WP_002191947.1 (68.0% identity, 78.0% similarity) Cry49Ab1 (21.9% identity, 33.7% similarity) APG00251 97 285 APG00233 (81.8% identity, 87.8% similarity) Cry 40, 45, 50, 55, 60, 65, 70, 75, 55, 60, 65, 70, 75, 80, 85, 90, 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 US20130227743A1_200 (37.8% identity, 50.2% similarity) AGA40058.1 (26.1% identity, 41.6% similarity) WP_002140071.1 (24.2% identity, 35.0% similarity) Cry3Ca1 (18.9% identity, 30.3% similarity) APG00255 98 286 APG00401 (94.1% identity, 96.9% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 US20130227743A1_200 (31.3% identity, 46.8% similarity) WP_017762616.1 (28.7% identity, 44.7% similarity) AGA40058.1 (28.5% identity, 44.6% similarity) Cry41Aa1 (19.3% identity, 28.9% similarity) APG00258 99 WP_020799009.1 (72.7% identity, 84.5% similarity) Cry6 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_037330331.1 (33.2% identity, 47.0% similarity) WP_040871902.1 (32.4% identity, 47.5% similarity) Cry6Ba1 (23.6% identity, 40.5% similarity) APG00261 100 287 WP_002187944.1 (76.1% identity, 85.5% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_001258160.1 (75.6% identity, 85.0% similarity) WP_001258161.1 (75.6% identity, 85.0% similarity) APG00454 (68.7% identity, 76.8% similarity) APG00242 (68.5% identity, 76.8% similarity) APG00223 (68.5% identity, 76.6% similarity) APG00118 (68.1% identity, 76.4% similarity) APG00065 (59.2% identity, 69.8% similarity) Cry35Aa2 (21.5% identity, 37.8% similarity) APG00262 101 288, 289 WP_002191947.1 (77.7% identity, 82.3% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 WP_002166959.1 (77.4% identity, 82.3% similarity) APG00925 (73.7% identity, 82.5% similarity) APG00369 (71.9% identity, 80.2% similarity) WP_000839920.1 (71.0% identity, 77.7% similarity) APG00065 (68.7% identity, 77.2% similarity) APG00212 (68.7% identity, 77.1% similarity) APG00249 (67.4% identity, 75.9% similarity) Cry49Aa1 (20.3% identity, 33.3% similarity) APG00263 102 WP_037330331.1 (32.9% identity, 51.0% similarity) Cry6 35, 40, 45, 50, 55, 60, 65, 70, 55, 60, 65, 70, 75, 80, 85, 90, 75, 80, 85, 90, 95, 96, 97, 98, 95, 96, 97, 98, 99 99 WP_040871901.1 (32.9% identity, 49.5% similarity) WP_040871902.1 (31.7% identity, 48.8% similarity) Cry6Ba1 (19.2% identity, 32.2% similarity) APG00264 103 290, 291 APG00307 (66.0% identity, 78.8% similarity) Cry6 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 APG00297 (64.5% identity, 73.6% similarity) WP_017762448.1 (64.4% identity, 73.9% similarity) WP_016110403.1 (29.0% identity, 49.5% similarity) WP_002140073.1 (28.9% identity, 49.1% similarity) WP_016083929.1 (26.8% identity, 47.6% similarity) APG00265 104 APG00291 (86.4% identity, 91.2% similarity) Cry6 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_001104853.1 (73.1% identity, 84.5% similarity) WP_002187785.1 (72.1% identity, 84.5% similarity) WP_001104975.1 (71.7% identity, 83.4% similarity) WP_016078425.1 (71.7% identity, 83.4% similarity) APG00280 (63.0% identity, 74.7% similarity) APG00720 (51.2% identity, 74.4% similarity) APG00381 (50.9% identity, 73.0% similarity) APG00266 105 292 APG00288 (55.5% identity, 69.9% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40058.1 (31.4% identity, 46.0% similarity) AGP17992.1 (27.9% identity, 39.9% similarity) WP_017762581.1 (25.8% identity, 37.3% similarity) WP_017762619.1 (24.9% identity, 36.6% similarity) APG00267 106 293, 294 APG00391 (93.5% identity, 96.1% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 50, 55, 60, 65, 70, 75, 80, 85, 70, 75, 80, 85, 90, 95, 96, 97, 90, 95, 96, 97, 98, 99 98, 99 US20130227743A1_200 (29.7% identity, 46.1% similarity) AGA40058.1 (28.1% identity, 44.7% similarity) WP_017762581.1 (25.9% identity, 39.9% similarity) AGP17992.1 (25.2% identity, 37.9% similarity) APG00270 107 WP_007973068.1 (77.4% identity, 90.3% similarity) Cry6 80, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00319 (75.8% identity, 89.8% similarity) WP_003379701.1 (43.2% identity, 62.2% similarity) WP_017683125.1 (43.2% identity, 62.2% similarity) WP_024677851.1 (43.0% identity, 62.0% similarity) APG00271 108 295 WP_018765648.1 (57.1% identity, 74.1% similarity) Cry6 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 WP_018780622.1 (56.7% identity, 74.1% similarity) WP_040119596.1 (56.7% identity, 74.1% similarity) WP_000790920.1 (56.5% identity, 74.6% similarity) APG00273 109 296 APG00033 (80.3% identity, 88.1% similarity) Vip 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00175 (78.0% identity, 85.7% similarity) APG00077 (77.1% identity, 84.9% similarity) APG00104 (72.1% identity, 82.1% similarity) KEZ80024.1 (69.1% identity, 79.7% similarity) APG00278 (68.9% identity, 78.0% similarity) APG00173 (68.8% identity, 78.1% similarity) US_7129212_B2-17 (23.7% identity, 41.2% similarity) US_8334431_B2-13 (23.6% identity, 41.2% similarity) Vip3Ca1 (23.0% identity, 37.3% similarity) APG00274 110 297, 298 WP_016098095.1 (93.1% identity, 97.1% similarity) Cry6 95, 96, 97, 98, 99 98, 99 WP_002064828.1 (77.4% identity, 87.7% similarity) WP_016103147.1 (77.4% identity, 87.7% similarity) WP_002126822.1 (77.1% identity, 87.7% similarity) APG00275 111 299 AGA40057.1 (26.0% identity, 30.3% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 35, 40, 45, 50, 55, 60, 65, 70, 70, 75, 80, 85, 90, 95, 96, 97, 75, 80, 85, 90, 95, 96, 97, 98, 98, 99 99 US20130227743A1_206 (13.3% identity, 20.5% similarity) AGA40058.1 (12.5% identity, 20.2% similarity) WP_017762619.1 (12.4% identity, 18.5% similarity) APG00278 112 113 APG00173 (94.5% identity, 96.8% similarity) Vip 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00077 (82.3% identity, 88.3% similarity) APG00175 (80.7% identity, 86.6% similarity) KEZ80024.1 (73.9% identity, 84.1% similarity) APG00273 (68.9% identity, 78.0% similarity) APG00104 (68.7% identity, 79.0% similarity) APG00033 (68.3% identity, 78.5% similarity) Vip3Ca3 (24.8% identity, 42.0% similarity) APG00279 114 300 WP_020799009.1 (33.1% identity, 48.6% similarity) Cry6 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 WP_012620778.1 (29.4% identity, 46.8% similarity) WP_012103144.1 (29.0% identity, 46.2% similarity) Cry6Ba1 (23.4% identity, 36.7% similarity) APG00280 115 APG00265 (63.0% identity, 74.7% similarity) Cry6 60, 65, 70, 75, 80, 85, 90, 95, 70, 75, 80, 85, 90, 95, 96, 97, 96, 97, 98, 99 98, 99 APG00291 (59.3% identity, 71.9% similarity) WP_001104853.1 (55.2% identity, 69.0% similarity) WP_002187785.1 (54.5% identity, 69.7% similarity) WP_001104975.1 (54.5% identity, 69.0% similarity) WP_016078425.1 (53.8% identity, 68.3% similarity) APG00282 116 301 APG00925 (69.9% identity, 75.6% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 APG00369 (66.1% identity, 72.7% similarity) APG00295 (61.3% identity, 68.1% similarity) WP_002191947.1 (61.0% identity, 70.1% similarity) WP_002166959.1 (60.7% identity, 70.1% similarity) APG00262 (59.8% identity, 66.6% similarity) WP_016107065.1 (59.0% identity, 69.4% similarity) APG00065 (57.9% identity, 65.8% similarity) Cry35Ab3 (21.5% identity, 36.3% similarity) APG00284 117 302 APG00731 (91.2% identity, 96.0% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_000143307.1 (89.3% identity, 94.4% similarity) WP_000143308.1 (89.0% identity, 93.8% similarity) APG00035 (88.7% identity, 95.4% similarity) APG00377 (86.6% identity, 92.5% similarity) APG00231 (86.3% identity, 92.2% similarity) APG00356 (85.8% identity, 92.5% similarity) US20130227743A1_6 (76.9% identity, 83.4% similarity) APG00011 (73.7% identity, 84.5% similarity) Cry35Ac2 (22.2% identity, 41.1% similarity) APG00285 118 303 WP_017762581.1 (30.9% identity, 45.1% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40057.1 (30.5% identity, 46.6% similarity) AGP17992.1 (30.5% identity, 43.9% similarity) WP_017762616.1 (30.4% identity, 45.2% similarity) APG00286 119 120, 304 WP_003204425.1 (53.7% identity, 74.3% similarity) Cry6 55, 60, 65, 70, 75, 80, 85, 90, 75, 80, 85, 90, 95, 96, 97, 98, 95, 96, 97, 98, 99 99 WP_018767550.1 (53.7% identity, 74.3% similarity) WP_018783608.1 (53.7% identity, 74.0% similarity) WP_003209431.1 (53.5% identity, 74.0% similarity) APG00287 121 305 APG00356 (98.4% identity, 98.7% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00377 (95.7% identity, 97.9% similarity) APG00231 (94.4% identity, 97.3% similarity) APG00035 (90.1% identity, 93.8% similarity) APG00731 (89.5% identity, 93.3% similarity) WP_000143307.1 (85.0% identity, 90.6% similarity) WP_000143308.1 (84.2% identity, 90.9% similarity) US20130227743A1_6 (78.8% identity, 83.4% similarity) APG00011 (76.9% identity, 84.7% similarity) Cry35Ab4 (22.5% identity, 39.1% similarity) APG00288 122 306 APG00266 (55.5% identity, 69.9% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40058.1 (31.7% identity, 47.3% similarity) AGP17992.1 (28.9% identity, 42.1% similarity) WP_017762581.1 (28.8% identity, 41.8% similarity) AGA40057.1 (26.9% identity, 39.9% similarity) APG00289 123 WP_017683125.1 (47.1% identity, 65.9% similarity) Cry6 50, 55, 60, 65, 70, 75, 80, 85, 70, 75, 80, 85, 90, 95, 96, 97, 90, 95, 96, 97, 98, 99 98, 99 WP_024677851.1 (47.1% identity, 65.7% similarity) WP_017701944.1 (46.9% identity, 65.9% similarity) WP_011105063.1 (46.9% identity, 65.4% similarity) APG00290 124 307 APG00422 (85.0% identity, 86.9% similarity) Cyt 30, 35, 40, 45, 50, 55, 60, 65, 40, 45, 50, 55, 60, 65, 70, 75, 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00698 (82.2% identity, 88.2% similarity) APG00647 (81.1% identity, 86.6% similarity) Cyt1Ba1 (25.8% identity, 39.2% similarity) APG00291 125 308 APG00265 (86.4% identity, 91.2% similarity) Cry6 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_001104853.1 (71.6% identity, 82.8% similarity) WP_001104975.1 (70.6% identity, 82.1% similarity) WP_002187785.1 (70.3% identity, 82.8% similarity) WP_016078425.1 (69.9% identity, 82.8% similarity) APG00280 (59.3% identity, 71.9% similarity) APG00381 (50.5% identity, 71.2% similarity) APG00720 (50.2% identity, 73.6% similarity) APG00292 126 309 US20130227743A1_200 (33.4% identity, 48.3% Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, similarity) 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40058.1 (26.5% identity, 42.3% similarity) AGP17992.1 (25.8% identity, 41.0% similarity) Cry42Aa1 (15.7% identity, 25.6% similarity) APG00294 127 310 WP_016102361.1 (51.7% identity, 70.6% similarity) Cry6 55, 60, 65, 70, 75, 80, 85, 90, 75, 80, 85, 90, 95, 96, 97, 98, 95, 96, 97, 98, 99 99 WP_002144067.1 (51.4% identity, 70.4% similarity) WP_002035071.1 (50.0% identity, 69.2% similarity) WP_040119237.1 (49.5% identity, 69.7% similarity) APG00295 128 311, 312 APG00308 (77.6% identity, 82.9% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 APG00385 (70.5% identity, 79.8% similarity) APG00392 (68.9% identity, 79.1% similarity) APG00767 (66.6% identity, 76.3% similarity) APG00065 (64.4% identity, 74.3% similarity) WP_033679178.1 (63.7% identity, 74.1% similarity) WP_000839920.1 (62.6% identity, 73.5% similarity) WP_002191947.1 (61.3% identity, 73.2% similarity) Cry49Aa1 (20.5% identity, 34.0% similarity) APG00297 129 313, 314 WP_017762448.1 (87.7% identity, 89.1% similarity) Cry6 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00264 (64.5% identity, 73.6% similarity) APG00307 (63.1% identity, 75.4% similarity) WP_033674645.1 (28.2% identity, 44.8% similarity) WP_000751842.1 (27.6% identity, 47.9% similarity) WP_000751829.1 (27.4% identity, 47.9% similarity) APG00298 130 315, 316 WP_016098094.1 (94.0% identity, 97.2% similarity) Cry6 95, 96, 97, 98, 99 98, 99 KFN03392.1 (77.7% identity, 87.3% similarity) WP_033674722.1 (74.4% identity, 86.8% similarity) WP_002135640.1 (73.1% identity, 85.8% similarity) APG00241 (52.0% identity, 71.0% similarity) APG00301 131 317 APG00353 (93.5% identity, 95.6% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00844 (91.1% identity, 94.8% similarity) APG00419 (87.9% identity, 90.4% similarity) APG00412 (87.5% identity, 91.3% similarity) APG00065 (77.7% identity, 84.4% similarity) WP_000839920.1 (73.8% identity, 83.0% similarity) WP_002166959.1 (68.8% identity, 79.4% similarity) WP_002114997.1 (67.9% identity, 78.8% similarity) Cry35Ac2 (22.5% identity, 37.8% similarity) APG00302 132 318, 319 AGA40058.1 (27.8% identity, 41.4% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US_2013_0227743_A1_178 (27.5% identity, 41.7% similarity) US20130227743A1_200 (26.9% identity, 44.0% similarity) Cry42Aa1 (17.9% identity, 29.2% similarity) APG00305 133 320 WP_017762447.1 (92.7% identity, 96.0% similarity) Cry6 95, 96, 97, 98, 99 97, 98, 99 WP_000162958.1 (44.7% identity, 60.9% similarity) WP_002012120.1 (44.7% identity, 60.7% similarity) WP_002118280.1 (44.7% identity, 60.7% similarity) APG00307 134 321, 322 APG00264 (66.0% identity, 78.8% similarity) Cry6 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_017762448.1 (63.9% identity, 76.3% similarity) APG00297 (63.1% identity, 75.4% similarity) WP_033674645.1 (26.9% identity, 46.1% similarity) KFM95248.1 (26.5% identity, 47.5% similarity) WP_000751870.1 (25.7% identity, 47.0% similarity) APG00308 135 323 APG00392 (82.9% identity, 91.2% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00767 (81.8% identity, 89.6% similarity) APG00295 (77.6% identity, 82.9% similarity) APG00385 (76.1% identity, 86.6% similarity) APG00065 (67.0% identity, 78.6% similarity) WP_033679178.1 (66.8% identity, 77.4% similarity) WP_000839920.1 (66.8% identity, 77.3% similarity) WP_002191947.1 (64.9% identity, 76.7% similarity) Cry35Ab1 (21.7% identity, 36.6% similarity) APG00314 136 324 WP_029439068.1 (72.5% identity, 81.9% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00415 (55.1% identity, 66.4% similarity) WP_029439066.1 (46.9% identity, 61.0% similarity) WP_002191947.1 (34.3% identity, 50.9% similarity) Cry35Ac2 (22.3% identity, 37.9% similarity) APG00315 137 APG00178 (87.7% identity, 91.6% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00727 (85.5% identity, 89.9% similarity) Cry49Ab1 (27.3% identity, 44.2% similarity) APG00317 138 WP_015096649.1 (68.9% identity, 84.4% similarity) Cry6 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00400 (68.7% identity, 84.6% similarity) APG00630 (67.2% identity, 83.6% similarity) WP_028939439.1 (45.1% identity, 63.9% similarity) WP_019823923.1 (32.5% identity, 51.3% similarity) WP_016779464.1 (31.5% identity, 51.4% similarity) APG00319 139 APG00270 (75.8% identity, 89.8% similarity) Cry6 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 WP_007973068.1 (71.2% identity, 86.8% similarity) WP_024691985.1 (41.9% identity, 62.8% similarity) WP_016569001.1 (41.7% identity, 62.8% similarity) WP_029571787.1 (41.7% identity, 62.8% similarity) APG00320 140 325 APG00913 (75.8% identity, 85.4% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00118 (72.2% identity, 81.7% similarity) APG00223 (70.8% identity, 81.7% similarity) APG00398 (68.3% identity, 77.3% similarity) WP_002114997.1 (67.6% identity, 75.7% similarity) WP_002187944.1 (61.9% identity, 72.6% similarity) US20130227743A1_10 (61.5% identity, 72.6% similarity) APG00065 (54.9% identity, 67.1% similarity) Cry35Ab4 (20.4% identity, 35.2% similarity) APG00340 141 326 AGP18023.1 (63.5% identity, 75.5% similarity) Cry 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 APG00151 (62.2% identity, 74.6% similarity) US20130227743A1_40 (56.7% identity, 65.9% similarity) US20130227743A1_48 (35.7% identity, 45.6% similarity) Cry4Ba4 (25.5% identity, 36.7% similarity) APG00342 142 WP_002144456.1 (89.8% identity, 95.8% similarity) Cry6 90, 95, 96, 97, 98, 99 96, 97, 98, 99 WP_002169783.1 (86.4% identity, 92.4% similarity) WP_016078427.1 (68.0% identity, 82.2% similarity) APG00024 (66.9% identity, 80.8% similarity) APG00180 (62.9% identity, 78.8% similarity) Cry6Ba1 (28.4% identity, 46.7% similarity) APG00349 143 APG00797 (90.2% identity, 94.8% similarity) Bin 45, 50, 55, 60, 65, 70, 75, 80, 65, 70, 75, 80, 85, 90, 95, 96, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_000143308.1 (43.0% identity, 61.6% similarity) WP_000143307.1 (42.5% identity, 61.6% similarity) US20130227743A1_6 (40.1% identity, 59.6% similarity) Cry35Ac1 (26.5% identity, 43.3% similarity) APG00353 144 327 APG00203 (95.3% identity, 96.6% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00301 (93.5% identity, 95.6% similarity) APG00844 (91.1% identity, 94.5% similarity) APG00412 (88.5% identity, 91.8% similarity) APG00065 (76.7% identity, 83.9% similarity) WP_000839920.1 (73.0% identity, 82.7% similarity) WP_002166959.1 (69.3% identity, 80.2% similarity) WP_002114997.1 (67.9% identity, 79.3% similarity) Cry35Ad2 (24.8% identity, 39.6% similarity) APG00355 145 146 WP_017762616.1 (23.5% identity, 39.5% similarity) Cry 25, 30, 35, 40, 45, 50, 55, 60, 40, 45, 50, 55, 60, 65, 70, 75, 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 AGA40057.1 (23.1% identity, 35.9% similarity) AGA40058.1 (23.0% identity, 37.4% similarity) WP_017762581.1 (22.5% identity, 36.8% similarity) APG00356 147 328 APG00287 (98.4% identity, 98.7% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00377 (97.1% identity, 98.9% similarity) APG00231 (95.4% identity, 98.1% similarity) APG00035 (91.2% identity, 94.6% similarity) APG00731 (90.6% identity, 94.4% similarity) WP_000143307.1 (86.1% identity, 91.7% similarity) WP_000143308.1 (84.7% identity, 91.2% similarity) US20130227743A1_6 (79.9% identity, 84.5% similarity) APG00011 (77.7% identity, 85.5% similarity) Cry35Ab4 (22.8% identity, 39.1% similarity) APG00368 148 329 APG00211 (98.7% identity, 99.2% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00011 (94.9% identity, 96.5% similarity) APG00377 (79.1% identity, 86.9% similarity) APG00356 (79.1% identity, 86.4% similarity) APG00231 (78.6% identity, 86.4% similarity) APG00035 (77.3% identity, 84.8% similarity) WP_000143307.1 (76.7% identity, 85.0% similarity) WP_000143308.1 (75.4% identity, 84.5% similarity) US20130227743A1_6 (69.3% identity, 78.6% similarity) Cry35Ac2 (22.3% identity, 38.6% similarity) APG00369 149 330 APG00925 (79.6% identity, 87.1% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 APG00262 (71.9% identity, 80.2% similarity) APG00282 (66.1% identity, 72.7% similarity) APG00913 (64.9% identity, 77.4% similarity) WP_002191947.1 (62.9% identity, 70.5% similarity) WP_002166959.1 (62.6% identity, 70.5% similarity) APG00065 (61.8% identity, 70.8% similarity) US20130227743A1_148 (59.6% identity, 67.6% similarity) Cry49Aa1 (19.1% identity, 33.5% similarity) APG00372 150 331 AGP17992.1 (36.7% identity, 46.7% similarity) Cry 40, 45, 50, 55, 60, 65, 70, 75, 50, 55, 60, 65, 70, 75, 80, 85, 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_017762581.1 (36.4% identity, 45.8% similarity) AGA40058.1 (32.6% identity, 48.1% similarity) WP_017762619.1 (30.3% identity, 44.2% similarity) APG00373 151 332 APG00413 (64.7% identity, 74.8% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_017154552.1 (64.2% identity, 75.9% similarity) APG00090 (62.8% identity, 78.5% similarity) KEZ80012.1 (62.5% identity, 73.5% similarity) APG00230 (61.4% identity, 75.2% similarity) APG00596 (61.2% identity, 74.0% similarity) APG00191 (59.8% identity, 72.8% similarity) APG00757 (57.2% identity, 71.8% similarity) WP_003308586.1 (48.6% identity, 62.0% similarity) Cry35Ab1 (21.5% identity, 33.3% similarity) APG00375 152 333 AGA40058.1 (43.6% identity, 59.3% similarity) Cry 45, 50, 55, 60, 65, 70, 75, 80, 60, 65, 70, 75, 80, 85, 90, 95, 85, 90, 95, 96, 97, 98, 99 96, 97, 98, 99 AGP17992.1 (35.7% identity, 50.5% similarity) WP_017762581.1 (34.7% identity, 48.2% similarity) AGA40057.1 (31.7% identity, 45.5% similarity) APG00376 153 334 APG00221 (92.5% identity, 95.1% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00237 (85.3% identity, 89.8% similarity) APG00353 (79.4% identity, 85.8% similarity) APG00412 (75.8% identity, 81.8% similarity) WP_002166959.1 (70.8% identity, 80.8% similarity) WP_002191947.1 (70.5% identity, 80.8% similarity) WP_000839920.1 (70.1% identity, 81.4% similarity) APG00065 (66.5% identity, 76.3% similarity) Cry35Ac1 (21.3% identity, 36.5% similarity) APG00377 154 335 APG00356 (97.1% identity, 98.9% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00231 (96.8% identity, 98.7% similarity) APG00287 (95.7% identity, 97.9% similarity) APG00731 (89.3% identity, 93.6% similarity) APG00035 (88.5% identity, 93.8% similarity) WP_000143307.1 (86.1% identity, 91.2% similarity) WP_000143308.1 (84.7% identity, 90.9% similarity) US20130227743A1_6 (79.6% identity, 83.9% similarity) APG00011 (77.7% identity, 86.1% similarity) Cry35Ab4 (22.5% identity, 40.6% similarity) APG00379 155 336, 337 APG00356 (86.9% identity, 93.3% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00287 (86.6% identity, 93.3% similarity) APG00231 (86.3% identity, 93.0% similarity) APG00377 (86.3% identity, 92.8% similarity) APG00035 (80.4% identity, 89.5% similarity) WP_000143308.1 (78.8% identity, 87.4% similarity) WP_000143307.1 (77.2% identity, 86.9% similarity) US20130227743A1_6 (72.7% identity, 81.0% similarity) APG00011 (71.6% identity, 82.6% similarity) Cry35Ab4 (25.2% identity, 42.6% similarity) APG00381 156 APG00397 (92.7% identity, 96.9% similarity) Cry6 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00720 (80.5% identity, 89.9% similarity) WP_001072414.1 (80.1% identity, 88.2% similarity) WP_002144454.1 (78.7% identity, 87.5% similarity) WP_002187783.1 (77.0% identity, 86.4% similarity) WP_002169785.1 (63.8% identity, 77.7% similarity) APG00265 (50.9% identity, 73.0% similarity) APG00291 (50.5% identity, 71.2% similarity) APG00385 157 338 APG00392 (80.2% identity, 89.9% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00767 (77.0% identity, 86.2% similarity) APG00407 (76.9% identity, 85.9% similarity) APG00308 (76.1% identity, 86.6% similarity) APG00065 (67.8% identity, 78.8% similarity) WP_002166959.1 (67.4% identity, 77.8% similarity) WP_002191947.1 (67.2% identity, 77.8% similarity) WP_000839920.1 (67.1% identity, 78.1% similarity) Cry35Ad2 (23.9% identity, 39.2% similarity) APG00386 158 WP_002167240.1 (88.1% identity, 91.4% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_016093722.1 (62.3% identity, 75.9% similarity) APG00215 (59.0% identity, 73.7% similarity) WP_002090518.1 (56.1% identity, 70.4% similarity) Cry35Ab1 (23.9% identity, 44.8% similarity) APG00388 159 WP_011898730.1 (43.7% identity, 60.0% similarity) Cyt 45, 50, 55, 60, 65, 70, 75, 80, 65, 70, 75, 80, 85, 90, 95, 96, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_005311350.1 (43.2% identity, 59.5% similarity) WP_017413134.1 (43.2% identity, 59.5% similarity) Cyt1Aa6 (30.5% identity, 51.6% similarity) APG00391 160 339 APG00267 (93.5% identity, 96.1% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 50, 55, 60, 65, 70, 75, 80, 85, 70, 75, 80, 85, 90, 95, 96, 97, 90, 95, 96, 97, 98, 99 98, 99 US20130227743A1_200 (29.7% identity, 46.5% similarity) AGA40058.1 (28.0% identity, 45.5% similarity) WP_017762581.1 (25.8% identity, 40.4% similarity) AGP17992.1 (25.5% identity, 39.0% similarity) APG00392 161 340 APG00308 (82.9% identity, 91.2% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00407 (82.1% identity, 88.3% similarity) APG00385 (80.2% identity, 89.9% similarity) APG00767 (79.0% identity, 88.2% similarity) APG00065 (73.4% identity, 83.7% similarity) US20130227743A1_146 (72.5% identity, 81.0% similarity) WP_000839920.1 (70.4% identity, 80.1% similarity) WP_002166959.1 (69.9% identity, 79.2% similarity) Cry49Aa1 (21.3% identity, 32.5% similarity) APG00395 162 341 AGA40057.1 (27.9% identity, 41.3% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 50, 55, 60, 65, 70, 75, 80, 85, 70, 75, 80, 85, 90, 95, 96, 97, 90, 95, 96, 97, 98, 99 98, 99 AGP17992.1 (27.5% identity, 40.9% similarity) WP_017762581.1 (27.4% identity, 41.4% similarity) AGA40058.1 (26.6% identity, 45.5% similarity) APG00396 163 342 AGA40057.1 (29.5% identity, 43.1% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 AGP17992.1 (28.6% identity, 42.9% similarity) WP_017762581.1 (28.1% identity, 40.8% similarity) US_2013_0227743_A1_178 (27.0% identity, 39.7% similarity) APG00397 164 APG00381 (92.7% identity, 96.9% similarity) Cry6 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00720 (79.4% identity, 89.2% similarity) WP_001072414.1 (79.1% identity, 87.1% similarity) WP_002144454.1 (77.4% identity, 86.4% similarity) WP_002187783.1 (76.0% identity, 86.1% similarity) WP_002169785.1 (63.4% identity, 77.4% similarity) APG00398 165 343, 344 APG00913 (85.9% identity, 88.4% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 80, 85, 90, 95, 96, 97, 98, 99 99 APG00118 (80.5% identity, 83.9% similarity) APG00223 (78.0% identity, 83.0% similarity) APG00454 (77.4% identity, 82.8% similarity) WP_002187944.1 (70.9% identity, 74.9% similarity) US20130227743A1_10 (70.2% identity, 75.5% similarity) WP_001258160.1 (70.0% identity, 75.3% similarity) APG00065 (56.6% identity, 67.6% similarity) Cry49Ab1 (18.7% identity, 31.1% similarity) APG00399 166 WP_023520621.1 (28.4% identity, 43.8% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_010720814.1 (25.1% identity, 36.9% similarity) WP_010829185.1 (15.6% identity, 25.4% similarity) Cry41Aa1 (13.1% identity, 21.9% similarity) APG00400 167 APG00630 (90.7% identity, 95.4% similarity) Cry6 95, 96, 97, 98, 99 96, 97, 98, 99 WP_015096649.1 (90.5% identity, 95.8% similarity) APG00317 (68.7% identity, 84.6% similarity) WP_028939439.1 (42.4% identity, 60.4% similarity) WP_019823923.1 (32.3% identity, 54.2% similarity) WP_016779464.1 (31.5% identity, 52.8% similarity) APG00401 168 345 APG00255 (94.1% identity, 96.9% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 US20130227743A1_200 (32.4% identity, 48.3% similarity) WP_017762616.1 (29.8% identity, 45.1% similarity) AGA40058.1 (29.6% identity, 44.4% similarity) Cry42Aa1 (18.9% identity, 30.2% similarity) APG00404 169 WP_017762616.1 (30.6% identity, 45.0% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40057.1 (28.3% identity, 41.0% similarity) AGA40058.1 (27.0% identity, 42.3% similarity) WP_017762581.1 (25.7% identity, 41.1% similarity) APG00407 170 346 APG00392 (82.1% identity, 88.3% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00065 (77.2% identity, 84.0% similarity) APG00385 (76.9% identity, 85.9% similarity) APG00132 (75.9% identity, 85.2% similarity) APG00249 (75.5% identity, 84.4% similarity) US20130227743A1_146 (71.0% identity, 79.7% similarity) WP_000839920.1 (70.8% identity, 80.8% similarity) WP_002166959.1 (70.5% identity, 79.2% similarity) Cry35Ad1 (21.1% identity, 35.4% similarity) APG00408 171 APG00067 (81.6% identity, 89.3% similarity) Mtx 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00559 (72.5% identity, 79.5% similarity) US20130227743A1_100 (69.9% identity, 80.9% similarity) WP_000963933.1 (69.7% identity, 81.0% similarity) APG00006 (68.3% identity, 80.1% similarity) APG00201 (68.3% identity, 78.1% similarity) APG00036 (67.4% identity, 78.1% similarity) APG00137 (66.2% identity, 78.0% similarity) APG00898 (65.9% identity, 79.5% similarity) APG00022 (64.2% identity, 77.2% similarity) US20130227743A1_60 (37.2% identity, 43.7% similarity) WP_026324166.1 (24.1% identity, 38.7% similarity) APG00409 172 347 WP_016576337.1 (42.8% identity, 53.8% similarity) Cyt 45, 50, 55, 60, 65, 70, 75, 80, 55, 60, 65, 70, 75, 80, 85, 90, 85, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_028812057.1 (42.4% identity, 52.3% similarity) WP_039629475.1 (42.0% identity, 54.2% similarity) Cyt2Aa2 (21.3% identity, 39.9% similarity) APG00412 173 348, 349 APG00844 (88.5% identity, 92.6% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00353 (88.5% identity, 91.8% similarity) APG00301 (87.5% identity, 91.3% similarity) APG00229 (84.9% identity, 89.1% similarity) APG00065 (75.7% identity, 84.4% similarity) WP_000839920.1 (70.4% identity, 81.1% similarity) WP_002166959.1 (67.6% identity, 78.0% similarity) WP_002191947.1 (67.3% identity, 78.0% similarity) Cry35Ab3 (23.4% identity, 37.7% similarity) APG00413 174 350 KEZ80012.1 (90.4% identity, 94.4% similarity) Bin 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00230 (81.0% identity, 88.9% similarity) WP_017154552.1 (73.5% identity, 83.6% similarity) APG00596 (69.6% identity, 82.2% similarity) APG00741 (69.0% identity, 79.3% similarity) APG00191 (66.2% identity, 79.2% similarity) APG00757 (64.8% identity, 74.1% similarity) APG00090 (63.7% identity, 79.3% similarity) US20130227743A1_156 (55.3% identity, 60.3% similarity) APG00060 (52.2% identity, 59.9% similarity) Cry35Ac2 (22.1% identity, 37.7% similarity) APG00415 175 351 WP_029439066.1 (68.8% identity, 81.5% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_029439068.1 (58.2% identity, 74.4% similarity) APG00314 (55.1% identity, 66.4% similarity) WP_033679178.1 (40.2% identity, 56.5% similarity) Cry35Ac2 (25.4% identity, 44.5% similarity) APG00419 176 352 APG00229 (92.6% identity, 94.1% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00844 (88.4% identity, 90.9% similarity) APG00353 (88.2% identity, 90.4% similarity) APG00301 (87.9% identity, 90.4% similarity) APG00065 (84.1% identity, 90.4% similarity) WP_000839920.1 (69.0% identity, 78.4% similarity) WP_002166959.1 (68.7% identity, 77.9% similarity) WP_002191947.1 (68.4% identity, 77.9% similarity) Cry35Ab5 (21.6% identity, 36.0% similarity) APG00422 177 APG00290 (85.0% identity, 86.9% similarity) Cyt 30, 35, 40, 45, 50, 55, 60, 65, 40, 45, 50, 55, 60, 65, 70, 75, 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00647 (78.0% identity, 83.3% similarity) APG00698 (77.5% identity, 83.0% similarity) Cyt1Aa6 (26.0% identity, 39.0% similarity) WP_000429377.1 (26.0% identity, 39.0% similarity) APG00427 178 353 APG00851 (88.4% identity, 93.1% similarity) Mtx 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_018673409.1 (80.7% identity, 89.3% similarity) WP_001039209.1 (68.4% identity, 81.0% similarity) US_8829279_B2-2 (55.7% identity, 70.6% similarity) US_8829279_B2-61 (55.7% identity, 70.6% similarity) APG00454 179 354 APG00242 (97.9% identity, 98.8% similarity) Bin 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00223 (97.7% identity, 98.4% similarity) APG00118 (90.3% identity, 94.9% similarity) US20130227743A1_10 (84.7% identity, 88.4% similarity) APG00913 (84.5% identity, 89.8% similarity) WP_001258160.1 (84.5% identity, 88.2% similarity) WP_001258161.1 (84.3% identity, 88.0% similarity) APG00065 (58.1% identity, 70.5% similarity) Cry35Ab4 (19.5% identity, 34.9% similarity) APG00456 180 WP_003203846.1 (94.1% identity, 95.6% similarity) Cyt 95, 96, 97, 98, 99 96, 97, 98, 99 WP_033798332.1 (94.1% identity, 95.6% similarity) WP_040119538.1 (94.1% identity, 95.6% similarity) Cyt2Ca1 (24.8% identity, 42.7% similarity) APG00459 181 355 APG00249 (89.0% identity, 92.8% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00132 (88.0% identity, 94.1% similarity) APG00065 (77.5% identity, 83.9% similarity) APG00419 (75.5% identity, 84.6% similarity) APG00407 (75.3% identity, 85.6% similarity) WP_000839920.1 (73.4% identity, 82.3% similarity) WP_002166959.1 (71.3% identity, 80.0% similarity) WP_002191947.1 (71.0% identity, 80.0% similarity) Cry35Ab4 (20.3% identity, 34.4% similarity) APG00461 182 APG00050 (64.2% identity, 74.1% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US_7692068_B2-2 (29.9% identity, 40.6% similarity) Cry49Aa1 (29.2% identity, 41.4% similarity) APG00462 183 WP_017413134.1 (85.4% identity, 92.1% similarity) Cyt 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_005311350.1 (85.0% identity, 91.7% similarity) AJE36343.1 (84.9% identity, 92.9% similarity) Cyt1Aa6 (33.3% identity, 48.0% similarity) APG00467 184 WP_017762581.1 (32.2% identity, 42.6% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 45, 50, 55, 60, 65, 70, 75, 80, 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 AGP17992.1 (31.9% identity, 43.9% similarity) US20130227743A1_184 (29.7% identity, 44.1% similarity) AGA40058.1 (27.5% identity, 43.0% similarity) APG00499 185 356 APG00532 (65.3% identity, 75.5% similarity) Bin 60, 65, 70, 75, 80, 85, 90, 95, 70, 75, 80, 85, 90, 95, 96, 97, 96, 97, 98, 99 98, 99 WP_003308586.1 (58.1% identity, 69.5% similarity) EJR24476.1 (53.9% identity, 59.5% similarity) KEZ80012.1 (48.5% identity, 60.7% similarity) Cry35Aa2 (19.4% identity, 34.1% similarity) APG00500 186 357 AGP17990.1 (63.6% identity, 75.3% similarity) Cry 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 APG00030 (57.5% identity, 64.9% similarity) APG00113 (56.6% identity, 65.3% similarity) APG00012 (56.3% identity, 58.2% similarity) APG00096 (51.8% identity, 60.6% similarity) APG00196 (51.7% identity, 62.1% similarity) AGP17989.1 (51.3% identity, 60.8% similarity) US_2013_0227743_A1_194 (50.8% identity, 58.2% similarity) Cry70Ba1 (23.3% identity, 37.3% similarity) APG00517 187 358 APG00227 (79.8% identity, 79.8% similarity) Mtx 45, 50, 55, 60, 65, 70, 75, 80, 50, 55, 60, 65, 70, 75, 80, 85, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_002166885.1 (40.6% identity, 49.8% similarity) US20130227743A1_110 (33.6% identity, 47.5% similarity) AGP17985.1 (28.6% identity, 42.5% similarity) US_8829279_B2-11 (28.4% identity, 41.1% similarity) APG00521 188 359 WP_000839061.1 (69.5% identity, 81.5% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US20130227743A1_146 (65.0% identity, 77.2% similarity) WP_002166959.1 (63.2% identity, 73.9% similarity) APG00243 (63.1% identity, 74.4% similarity) APG00065 (62.9% identity, 75.4% similarity) APG00132 (62.9% identity, 74.5% similarity) APG00249 (62.4% identity, 73.0% similarity) APG00212 (59.4% identity, 71.7% similarity) Cry35Aa1 (22.3% identity, 38.1% similarity) APG00532 189 APG00499 (65.3% identity, 75.5% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_003308586.1 (64.7% identity, 76.5% similarity) WP_017154552.1 (52.3% identity, 66.7% similarity) KEZ80012.1 (51.7% identity, 65.8% similarity) APG00230 (50.6% identity, 66.0% similarity) Cry35Aa2 (22.6% identity, 38.5% similarity) APG00550 190 WP_000727408.1 (41.4% identity, 56.0% similarity) Bin 45, 50, 55, 60, 65, 70, 75, 80, 60, 65, 70, 75, 80, 85, 90, 95, 85, 90, 95, 96, 97, 98, 99 96, 97, 98, 99 WP_000727409.1 (41.4% identity, 56.0% similarity) WP_016110923.1 (41.4% identity, 56.0% similarity) Cry35Ad2 (22.3% identity, 41.8% similarity) APG00559 191 360, 361 APG00067 (75.5% identity, 81.2% similarity) Mtx 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00408 (72.5% identity, 79.5% similarity) WP_000963933.1 (66.0% identity, 76.8% similarity) US20130227743A1_100 (65.6% identity, 75.6% similarity) APG00006 (63.6% identity, 74.4% similarity) APG00137 (63.5% identity, 75.7% similarity) APG00036 (63.2% identity, 73.6% similarity) APG00201 (63.2% identity, 73.0% similarity) APG00898 (62.8% identity, 73.8% similarity) APG00022 (60.8% identity, 73.8% similarity) US20130227743A1_60 (33.4% identity, 40.0% similarity) WP_037756193.1 (24.6% identity, 35.2% similarity) APG00571 192 362 APG00377 (76.5% identity, 84.7% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00356 (76.5% identity, 83.6% similarity) APG00731 (75.7% identity, 80.7% similarity) APG00035 (74.7% identity, 81.5% similarity) APG00284 (74.0% identity, 81.7% similarity) WP_000143307.1 (72.9% identity, 79.1% similarity) WP_000143308.1 (72.1% identity, 80.4% similarity) US20130227743A1_6 (65.8% identity, 73.9% similarity) APG00011 (64.4% identity, 73.5% similarity) Cry35Ac2 (24.8% identity, 42.9% similarity) APG00590 193 363 AGA40045.1 (73.7% identity, 82.6% similarity) Mtx 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00146 (68.1% identity, 79.8% similarity) WP_000794514.1 (67.8% identity, 80.4% similarity) US20130227743A1_102 (61.7% identity, 74.8% similarity) WP_036654376.1 (44.3% identity, 59.9% similarity) APG00591 194 WP_033679178.1 (48.6% identity, 59.9% similarity) Bin 50, 55, 60, 65, 70, 75, 80, 85, 65, 70, 75, 80, 85, 90, 95, 96, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_002166959.1 (48.5% identity, 61.2% similarity) WP_002191947.1 (48.2% identity, 61.2% similarity) Cry35Ac2 (27.3% identity, 45.9% similarity) APG00592 195 364, 365 APG00212 (89.8% identity, 94.2% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00798 (86.2% identity, 91.7% similarity) APG00619 (84.7% identity, 89.8% similarity) APG00600 (83.9% identity, 90.0% similarity) US20130227743A1_146 (74.9% identity, 82.6% similarity) APG00065 (68.9% identity, 77.2% similarity) WP_002166959.1 (67.1% identity, 76.8% similarity) WP_002191947.1 (66.8% identity, 76.8% similarity) Cry49Aa1 (22.1% identity, 35.4% similarity) APG00596 196 366 WP_017154552.1 (88.2% identity, 93.7% similarity) Bin 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 APG00741 (71.7% identity, 81.1% similarity) APG00413 (69.6% identity, 82.2% similarity) KEZ80012.1 (69.2% identity, 80.5% similarity) APG00230 (69.1% identity, 80.9% similarity) APG00757 (66.7% identity, 76.0% similarity) APG00191 (60.6% identity, 75.7% similarity) APG00090 (57.7% identity, 74.3% similarity) APG00060 (56.5% identity, 61.5% similarity) US20130227743A1_156 (55.1% identity, 61.9% similarity) Cry49Aa1 (19.3% identity, 30.8% similarity) APG00600 197 367 APG00212 (87.1% identity, 92.0% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00619 (85.6% identity, 93.3% similarity) APG00798 (84.7% identity, 90.8% similarity) APG00592 (83.9% identity, 90.0% similarity) US20130227743A1_146 (73.5% identity, 82.4% similarity) WP_002166959.1 (68.5% identity, 78.2% similarity) WP_002191947.1 (68.2% identity, 78.2% similarity) APG00065 (63.3% identity, 75.2% similarity) Cry35Ab3 (22.3% identity, 37.0% similarity) APG00603 198 WP_017762616.1 (32.7% identity, 49.5% similarity) Cry 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 AGA40058.1 (28.4% identity, 41.3% similarity) US20130227743A1_206 (25.2% identity, 39.8% similarity) WP_017762619.1 (24.6% identity, 40.7% similarity) APG00619 199 368 APG00798 (88.6% identity, 92.7% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00212 (86.2% identity, 92.0% similarity) APG00600 (85.6% identity, 93.3% similarity) APG00592 (84.7% identity, 89.8% similarity) US20130227743A1_146 (73.6% identity, 82.3% similarity) WP_002166959.1 (68.8% identity, 78.6% similarity) WP_002191947.1 (68.6% identity, 78.6% similarity) APG00065 (65.6% identity, 75.5% similarity) Cry35Ab4 (20.6% identity, 36.5% similarity) APG00630 200 APG00400 (90.7% identity, 95.4% similarity) Cry6 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_015096649.1 (87.5% identity, 93.9% similarity) APG00317 (67.2% identity, 83.6% similarity) WP_028939439.1 (41.9% identity, 59.9% similarity) WP_019823923.1 (32.4% identity, 52.1% similarity) WP_016779464.1 (30.4% identity, 52.1% similarity) APG00635 201 369, 370, 371 APG00020 (81.9% identity, 86.7% similarity) Mtx 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_002166885.1 (79.8% identity, 85.4% similarity) APG00227 (55.0% identity, 66.2% similarity) US20130227743A1_110 (44.0% identity, 60.2% similarity) AGP17985.1 (36.1% identity, 51.4% similarity) AGP17983.1 (34.9% identity, 52.1% similarity) APG00642 202 203 WP_029439068.1 (34.5% identity, 48.8% similarity) Bin 35, 40, 45, 50, 55, 60, 65, 70, 50, 55, 60, 65, 70, 75, 80, 85, 75, 80, 85, 90, 95, 96, 97, 98, 90, 95, 96, 97, 98, 99 99 WP_033679178.1 (33.9% identity, 47.6% similarity) R8EAW2_BACCE (32.6% identity, 47.2% similarity) Cry35Aa1 (22.1% identity, 36.9% similarity) APG00647 204 372 APG00698 (85.4% identity, 91.2% similarity) Cyt 25, 30, 35, 40, 45, 50, 55, 60, 40, 45, 50, 55, 60, 65, 70, 75, 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 APG00290 (81.1% identity, 86.6% similarity) APG00422 (78.0% identity, 83.3% similarity) GAJ51067.1 (24.9% identity, 35.1% similarity) Cyt1Ba1 (24.6% identity, 38.6% similarity) APG00652 205 373 WP_026592789.1 (50.4% identity, 68.7% similarity) Cry6 55, 60, 65, 70, 75, 80, 85, 90, 70, 75, 80, 85, 90, 95, 96, 97, 95, 96, 97, 98, 99 98, 99 WP_002175239.1 (49.9% identity, 69.7% similarity) WP_000591964.1 (49.3% identity, 69.5% similarity) WP_000591974.1 (49.3% identity, 69.5% similarity) APG00698 206 374 APG00647 (85.4% identity, 91.2% similarity) Cyt 25, 30, 35, 40, 45, 50, 55, 60, 40, 45, 50, 55, 60, 65, 70, 75, 65, 70, 75, 80, 85, 90, 95, 96, 80, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 APG00290 (82.2% identity, 88.2% similarity) APG00422 (77.5% identity, 83.0% similarity) Cyt1Ba1 (24.5% identity, 38.3% similarity) APG00700 207 APG00095 (97.9% identity, 98.2% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 Cry49Ab1 (28.7% identity, 40.9% similarity) APG00720 208 WP_001072414.1 (88.5% identity, 94.1% similarity) Cry6 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_002187783.1 (87.1% identity, 93.7% similarity) WP_002144454.1 (86.4% identity, 94.1% similarity) APG00381 (80.5% identity, 89.9% similarity) APG00397 (79.4% identity, 89.2% similarity) WP_002169785.1 (67.0% identity, 79.4% similarity) APG00265 (51.2% identity, 74.4% similarity) APG00291 (50.2% identity, 73.6% similarity) APG00723 209 375 WP_017762581.1 (28.9% identity, 41.0% similarity) Cry 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 AGA40057.1 (28.7% identity, 42.2% similarity) AGA40058.1 (27.8% identity, 40.4% similarity) AGP17992.1 (27.8% identity, 40.0% similarity) APG00727 210 APG00178 (90.7% identity, 93.6% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00315 (85.5% identity, 89.9% similarity) Cry49Aa1 (26.8% identity, 41.9% similarity) APG00731 211 376 APG00035 (95.4% identity, 98.4% similarity) Bin 95, 96, 97, 98, 99 98, 99 WP_000143307.1 (93.8% identity, 97.1% similarity) APG00284 (91.2% identity, 96.0% similarity) APG00356 (90.6% identity, 94.4% similarity) WP_000143308.1 (90.1% identity, 96.0% similarity) APG00287 (89.5% identity, 93.3% similarity) APG00231 (89.3% identity, 93.6% similarity) US20130227743A1_6 (82.6% identity, 86.6% similarity) APG00011 (76.4% identity, 85.3% similarity) Cry35Ac2 (21.1% identity, 40.5% similarity) APG00732 212 WP_016084067.1 (45.6% identity, 53.2% similarity) BIN 50, 55, 60, 65, 70, 75, 80, 85, 55, 60, 65, 70, 75, 80, 85, 90, 90, 95, 96, 97, 98, 99 95, 96, 97, 98, 99 WP_000836979.1 (45.4% identity, 53.6% similarity) WP_016123021.1 (36.1% identity, 47.0% similarity) Cry73Aa (16.7% identity, 26.2% similarity) APG00741 213 APG00757 (91.4% identity, 92.2% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 WP_017154552.1 (73.7% identity, 82.5% similarity) APG00060 (72.2% identity, 72.5% similarity) APG00596 (71.7% identity, 81.1% similarity) APG00413 (69.0% identity, 79.3% similarity) KEZ80012.1 (66.7% identity, 76.9% similarity) APG00230 (65.8% identity, 78.2% similarity) APG00090 (63.0% identity, 76.1% similarity) APG00191 (62.6% identity, 76.4% similarity) WP_003308586.1 (49.5% identity, 62.4% similarity) Cry49Aa1 (22.4% identity, 34.6% similarity) APG00748 214 377 WP_002016877.1 (43.1% identity, 61.8% similarity) Bin 45, 50, 55, 60, 65, 70, 75, 80, 65, 70, 75, 80, 85, 90, 95, 96, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_016097060.1 (42.0% identity, 59.4% similarity) WP_002167240.1 (40.5% identity, 56.5% similarity) Cry35Ad1 (22.3% identity, 37.4% similarity) APG00757 215 378 APG00741 (91.4% identity, 92.2% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00060 (69.4% identity, 69.4% similarity) WP_017154552.1 (68.8% identity, 77.3% similarity) APG00596 (66.7% identity, 76.0% similarity) APG00413 (64.8% identity, 74.1% similarity) KEZ80012.1 (63.2% identity, 73.1% similarity) APG00230 (62.3% identity, 73.8% similarity) APG00090 (59.0% identity, 72.0% similarity) APG00191 (58.6% identity, 71.7% similarity) WP_003308586.1 (49.8% identity, 63.6% similarity) Cry49Aa1 (21.1% identity, 32.6% similarity) APG00767 216 379 APG00308 (81.8% identity, 89.6% similarity) Bin 70, 75, 80, 85, 90, 95, 96, 97, 80, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00392 (79.0% identity, 88.2% similarity) APG00385 (77.0% identity, 86.2% similarity) APG00407 (71.4% identity, 81.9% similarity) WP_000839920.1 (65.0% identity, 76.3% similarity) WP_002166959.1 (63.1% identity, 75.2% similarity) WP_002191947.1 (62.9% identity, 75.2% similarity) APG00065 (62.4% identity, 76.3% similarity) Cry35Ab1 (20.9% identity, 36.5% similarity) APG00768 217 380 WP_000875422.1 (67.7% identity, 81.0% similarity) Mtx 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 US20130227743A1_58 (67.1% identity, 81.5% similarity) WP_000875423.1 (67.1% identity, 80.7% similarity) WP_000797173.1 (66.7% identity, 81.0% similarity) APG00055 (65.0% identity, 80.6% similarity) APG00174 (61.7% identity, 75.0% similarity) APG00064 (57.9% identity, 73.5% similarity) APG00797 218 APG00349 (90.2% identity, 94.8% similarity) Bin 45, 50, 55, 60, 65, 70, 75, 80, 65, 70, 75, 80, 85, 90, 95, 96, 85, 90, 95, 96, 97, 98, 99 97, 98, 99 WP_000143308.1 (43.6% identity, 61.7% similarity) WP_000143307.1 (42.8% identity, 61.7% similarity) US20130227743A1_6 (40.2% identity, 59.8% similarity) Cry35Ac1 (27.1% identity, 43.8% similarity) APG00798 219 381, 382 APG00212 (92.7% identity, 96.8% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00619 (88.6% identity, 92.7% similarity) APG00592 (86.2% identity, 91.7% similarity) APG00600 (84.7% identity, 90.8% similarity) US20130227743A1_146 (71.8% identity, 80.3% similarity) APG00065 (68.0% identity, 77.6% similarity) WP_002166959.1 (67.1% identity, 77.1% similarity) WP_002191947.1 (66.9% identity, 77.1% similarity) Cry35Ab4 (22.1% identity, 35.7% similarity) APG00830 220 383, 384 WP_033678680.1 (58.7% identity, 74.6% similarity) Cry6 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 WP_002120795.1 (58.7% identity, 73.7% similarity) WP_002161211.1 (58.7% identity, 73.4% similarity) WP_002199893.1 (58.5% identity, 73.7% similarity) APG00844 221 385 APG00353 (91.1% identity, 94.5% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 85, 90, 95, 96, 97, 98, 99 99 APG00412 (88.5% identity, 92.6% similarity) APG00419 (88.4% identity, 90.9% similarity) APG00229 (86.4% identity, 89.1% similarity) APG00065 (78.7% identity, 86.6% similarity) WP_000839920.1 (72.5% identity, 82.4% similarity) WP_002166959.1 (70.1% identity, 80.7% similarity) WP_002191947.1 (69.8% identity, 80.7% similarity) Cry35Ac2 (23.7% identity, 40.4% similarity) APG00851 222 386 APG00427 (88.4% identity, 93.1% similarity) Mtx 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 WP_018673409.1 (81.6% identity, 89.0% similarity) WP_001039209.1 (67.2% identity, 78.8% similarity) US_8829279_B2-61 (56.3% identity, 69.3% similarity) US_8829279_B2-2 (55.6% identity, 68.6% similarity) APG00862 223 WP_002167240.1 (27.4% identity, 40.1% similarity) Bin 30, 35, 40, 45, 50, 55, 60, 65, 45, 50, 55, 60, 65, 70, 75, 80, 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 WP_016093722.1 (27.3% identity, 42.1% similarity) WP_002016877.1 (25.7% identity, 38.9% similarity) Cry35Aa2 (19.4% identity, 31.1% similarity) APG00898 224 387 APG00201 (74.3% identity, 85.2% similarity) Mtx 70, 75, 80, 85, 90, 95, 96, 97, 85, 90, 95, 96, 97, 98, 99 98, 99 APG00036 (70.4% identity, 82.2% similarity) APG00006 (69.6% identity, 82.7% similarity) APG00137 (69.2% identity, 80.0% similarity) WP_000963933.1 (69.0% identity, 81.5% similarity) US20130227743A1_100 (68.8% identity, 80.6% similarity) APG00022 (68.2% identity, 79.2% similarity) APG00067 (67.9% identity, 80.7% similarity) APG00155 (67.1% identity, 79.3% similarity) APG00107 (64.8% identity, 78.8% similarity) US20130227743A1_60 (41.5% identity, 47.4% similarity) WP_026324166.1 (25.7% identity, 39.5% similarity) APG00907 225 388 AGA40061.1 (59.2% identity, 73.5% similarity) Mtx 60, 65, 70, 75, 80, 85, 90, 95, 75, 80, 85, 90, 95, 96, 97, 98, 96, 97, 98, 99 99 APG00112 (57.7% identity, 69.5% similarity) AGA40062.1 (54.2% identity, 69.3% similarity) Cry64Aa1 (38.3% identity, 52.6% similarity) APG00913 226 389 APG00118 (90.9% identity, 92.7% similarity) Bin 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 APG00398 (85.9% identity, 88.4% similarity) APG00223 (85.1% identity, 90.0% similarity) APG00454 (84.5% identity, 89.8% similarity) WP_002187944.1 (77.8% identity, 81.8% similarity) US20130227743A1_10 (75.8% identity, 81.6% similarity) WP_001258160.1 (75.6% identity, 81.4% similarity) APG00065 (60.4% identity, 72.8% similarity) Cry35Ac2 (18.9% identity, 33.3% similarity) APG00925 227 390 APG00369 (79.6% identity, 87.1% similarity) Bin 65, 70, 75, 80, 85, 90, 95, 96, 75, 80, 85, 90, 95, 96, 97, 98, 97, 98, 99 99 APG00262 (73.7% identity, 82.5% similarity) APG00282 (69.9% identity, 75.6% similarity) WP_016107065.1 (63.4% identity, 72.0% similarity) APG00065 (63.2% identity, 71.8% similarity) WP_002191947.1 (61.7% identity, 71.3% similarity) US20130227743A1_148 (61.7% identity, 70.2% similarity) APG00398 (60.2% identity, 71.9% similarity) Cry49Aa1 (19.5% identity, 34.2% similarity) APG00568 391 392 APG00356 (97.6% identity, 99.2% similarity) Bin 95, 96, 97, 98, 99 98, 99 APG00287 (96.2% identity, 98.1% similarity) APG00231 (95.7% identity, 95.7% similarity) APG00377 (95.4% identity, 98.7% similarity) WP_050845516.1 (93.8% identity, 97.9% similarity) AEX56523.1 (93.3% identity, 97.6% similarity) APG00035 (89.5% identity, 94.1% similarity) WP_000143307.1 (86.9% identity, 92.5% similarity) WP_000143308.1 (84.5% identity, 91.4% similarity) US_2013_0227743_A1-6 (78.6% identity, 84.2% similarity) APG00011 (77.7% identity, 85.5% similarity) APG00716 393 394 APG00376 (98.4% identity, 98.5% similarity) Bin 75, 80, 85, 90, 95, 96, 97, 98, 99 85, 90, 95, 96, 97, 98, 99 APG00221 (92.7% identity, 95.1% similarity) APG00237 (85.8% identity, 90.1% similarity) APG00353 (79.3% identity, 86.0% similarity) WP_000839920.1 (70.3% identity, 81.4% similarity) WP_002166959.1 (70.2% identity, 79.9% similarity) WP_002191947.1 (70.0% identity, 79.9% similarity) US_2013_0227743_A1-146 (67.2% identity, 77.0% similarity) APG00065 (66.2% identity, 76.5% similarity) APG00736 395 396 APG00647 (72.9% identity, 79.1% similarity) Cyt 70, 75, 80, 85, 90, 95, 96, 97, 98, 99 80, 85, 90, 95, 96, 97, 98, 99 APG00290 (71.8% identity, 79.5% similarity) APG00698 (70.8% identity, 78.4% similarity) APG00422 (69.9% identity, 75.8% similarity) WP_058323095.1 (69.9% identity, 75.8% similarity) APG00930 397 APG01245 (97.0% identity, 98.3% similarity) Mtx 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00107 (93.9% identity, 96.6% similarity) APG00155 (76.4% identity, 86.5% similarity) WP_000963933.1 (76.4% identity, 87.2% similarity) US_2013_0227743_A1-100 (76.0% identity, 86.5% similarity) APG00201 (74.3% identity, 82.9% similarity) APG00006 (73.9% identity, 82.7% similarity) APG00067 (70.8% identity, 83.9% similarity) APG00036 (70.8% identity, 80.4% similarity) APG00022 (69.4% identity, 80.7% similarity) APG01245 398 APG00930 (97.0% identity, 98.3% similarity) Mtx 80, 85, 90, 95, 96, 97, 98, 99 90, 95, 96, 97, 98, 99 APG00107 (93.2% identity, 96.6% similarity) US_2013_0227743_A1-100 (77.4% identity, 86.9% similarity) WP_000963933.1 (76.7% identity, 86.8% similarity) APG00155 (75.7% identity, 85.8% similarity) APG00201 (74.3% identity, 81.9% similarity) APG00006 (74.1% identity, 83.9% similarity) APG00067 (71.1% identity, 83.6% similarity) APG00036 (70.0% identity, 80.3% similarity) APG00022 (69.4% identity, 80.4% similarity)

i. Classes of Pesticidal Proteins

The pesticidal proteins provided herein and the nucleotide sequences encoding them are useful in methods for impacting pests. That is, the compositions and methods of the invention find use in agriculture for controlling or killing pests, including pests of many crop plants. The pesticidal proteins provided herein are toxin proteins from bacteria and exhibit activity against certain pests. The pesticidal proteins are from several classes of toxins including Cry, Cyt, BIN, and Mtx toxins. See, for example, Table 1 for the specific protein classifications of the various SEQ ID NOS provided herein. In addition, reference is made throughout this disclosure to Pfam database entries. The Pfam database is a database of protein families, each represented by multiple sequence alignments and a profile hidden Markov model. Finn et al. (2014) Nucl. Acid Res. Database Issue 42:D222-D230.

Bacillus thuringiensis (Bt) is a gram-positive bacterium that produces insecticidal proteins as crystal inclusions during its sporulation phase of growth. The proteinaceous inclusions of Bacillus thuringiensis (Bt) are called crystal proteins or 6-endotoxins (or Cry proteins), which are toxic to members of the class Insecta and other invertebrates. Similarly, Cyt proteins are parasporal inclusion proteins from Bt that exhibits hemolytic (cytolytic) activity or has obvious sequence similarity to a known Cyt protein. These toxins are highly specific to their target organism, and are innocuous to humans, vertebrates, and plants.

The structure of the Cry toxins reveals five conserved amino acid blocks, concentrated mainly in the center of the domain or at the junction between the domains. The Cry toxin consists of three domains, each with a specific function. Domain I is a seven α-helix bundle in which a central helix is completely surrounded by six outer helices. This domain is implicated in channel formation in the membrane. Domain II appears as a triangular column of three anti-parallel β-sheets, which are similar to antigen-binding regions of immunoglobulins. Domain III contains anti-parallel β-strands in a β sandwich form. The N-terminal part of the toxin protein is responsible for its toxicity and specificity and contains five conserved regions. The C-terminal part is usually highly conserved and probably responsible for crystal formation. See, for example, U.S. Pat. No. 8,878,007.

Strains of B. thuringiensis show a wide range of specificity against different insect orders (Lepidoptera, Diptera, Coleoptera, Hymenoptera, Homoptera, Phthiraptera or Mallophaga, and Acari) and other invertebrates (Nemathelminthes, Platyhelminthes, and Sarocomastebrates). The Cry proteins have been classified into groups based on toxicity to various insect and invertebrate groups. Generally, Cry I demonstrates toxicity to lepidopterans, Cry II to lepidopterans and dipterans, CryIII to coleopterans, Cry IV to dipterans, and Cry V and Cry VI to nematodes. New Cry proteins can be identified and assigned to a Cry group based on amino acid identity. See, for example, Bravo, A. (1997) J. of Bacteriol. 179:2793-2801; Bravo et al. (2013) Microb. Biotechnol. 6:17-26, herein incorporated by reference.

Over 750 different cry gene sequences have been classified into 73 groups (Cry1-Cry73), with new members of this gene family continuing to be discovered. The cry gene family consists of several phylogentically non-related protein families that may have different modes of action: the family of three-domain Cry toxins, the family of mosquitocidal Cry toxins, the family of the binary-like toxins, and the Cyt family of toxins (Bravo et al., 2005). Some Bt strains produce additional insecticidal toxins, the VIP toxins. See, also, Cohen et al. (2011) J. Mol. Biol. 413:4-814; the Bacillus thuringiensis Toxin Nomenclature database maintained by Dr. Neil Crickmore (Bacillus thruingiensis Lab, School of Life Sciences, University of Sussex) of the Bacillus thuringiensis delta-endotoxin nomenclature committee; Crickmore et al. (1988) Microbiol. Mol. Biol. Rev. 62: 807-813; Gill et al. (1992) Ann. Rev. Entomol. 37: 807-636; Goldbert et al. (1997) Appl. Environ. Microbiol. 63:2716-2712; Knowles et al. (1992) Proc. R. Soc. Ser. B. 248: 1-7; Koni et al. (1994) Microbiology 140: 1869-1880; Lailak et al. (2013) Biochem. Biophys. Res. Commun. 435: 216-221; Lopez-Diaz et al. (2013) Environ. Microbiol. 15: 3030-3039; Perez et al. (2007) Cell. Microbiol. 9: 2931-2937; Promdonkoy et al. (2003) Biochem. J. 374: 255-259; Rigden (2009) FEBS Lett. 583: 1555-1560; Schnepf et al. (1998)Microbiol. Mol. Biol. Rev. 62: 775-806; Soberon et al. (2013) Peptides 41: 87-93; Thiery et al. (1998) J. Am. Mosq. Control Assoc. 14: 472-476; Thomas et al. (1983) FEBS Lett. 154: 362-368; Wirth et al. (1997) Proc. Natl. Acad. Sci. U.S.A. 94: 10536-10540; Wirth et al (2005) Appl. Environ. Microbiol. 71: 185-189; and, Zhang et al. (2006) Biosci. Biotechnol. Biochem. 70: 2199-2204; each of which is herein incorporated by reference in their entirety.

Cyt designates a parasporal crystal inclusion protein from Bacillus thuringiensis with cytolytic activity, or a protein with sequence similarity to a known Cyt protein. (Crickmore et al. (1998) Microbial. Mol. Biol. Rev. 62: 807-813). The gene is denoted by cyt. These proteins are different in structure and activity from Cry proteins (Gill et al. (1992) Annu. Rev. Entomol. 37: 615-636). The Cyt toxins were first discovered in B. thuringiensis subspecies israelensis (Goldberg et al. (1977) Mosq. News. 37: 355-358). There are 3 Cyt toxin families including 11 holotype toxins in the current nomenclature (Crickmore et al. (2014) Bacillus thuringiensis toxin nomenclature found on the world wide web atlifesci.sussex.ac.uk/home/Neil_Crickmore/Bt/). The majority of the B. thuringiensis isolates with cyt genes show activity against dipteran insects (particularly mosquitoes and black flies), but there are also cyt genes that have been described in B. thuringiensis strains targeting lepidopteran or coleopteran insects (Guerchicoff et al. (1997) Appl. Environ. Microbiol. 63: 2716-2721).

The structure of Cyt2A, solved by X-ray crystallography, shows a single domain where two outer layers of α-helix wrap around a mixed β-sheet. Further available crystal structures of Cyt toxins support a conserved α-β structural model with two α-helix hairpins flanking a β-sheet core containing seven to eight β-strands. (Cohen et al. (2011) J. Mol. Biol. 413: 80 4-814) Mutagenic studies identified β-sheet residues as critical for toxicity, while mutations in the helical domains did not affect toxicity (Adang et al.; Diversity of Bacillus thuringiensis Crystal Toxins and Mechanism of Action. In: T. S. Dhadialla and S. S. Gill, eds, Advances in Insect Physiology, Vol. 47, Oxford: Academic Press, 2014, pp. 39-87.) The representative domain of the Cyt toxin is a δ-endotoxin, Bac_thur_toxin (Pfam PF01338).

There are multiple proposed models for the mode of action of Cyt toxins, and it is still an area of active investigation. Some Cyt proteins (Cyt1A) have been shown to require the presence of accessory proteins for crystallization. Cyt1A and Cyt2A protoxins are processed by digestive proteases at the same sites in the N- and C-termini to a stable toxin core. Cyt toxins then interact with non-saturated membrane lipids, such as phosphatidylcholine, phosphatidylethanolamine, and sphingomyelin. For Cyt toxins, pore-formation and detergent-like membrane disruption have been proposed as non-exclusive mechanisms; and it is generally accepted that both may occur depending on toxin concentration, with lower concentrations favoring oligomeric pores and higher concentrations leading to membrane breaks. (Butko (2003) Appl. Environ. Microbiol. 69: 2415-2422) In the pore-formation model, the Cyt toxin binds to the cell membrane, inducing the formation of cation-selective channels in the membrane vesicles leading to colloid-osmotic lysis of the cell. (Knowles et al. (1989) FEBS Lett. 244: 259-262; Knowles et al. (1992) Proc. R. Soc. Ser. B. 248: 1-7 and Promdonkoy et al. (2003) Biochem. J. 374: 255-259). In the detergent model, there is a nonspecific aggregation of the toxin on the surface of the lipid bilayer leading to membrane disassembly and cell death. (Butko (2003) supra; Manceva et al. (2005) Biochem. 44: 589-597).

Multiple studies have shown synergistic activity between Cyt toxins and other B. thuringiensis toxins, particularly the Cry, Bin, and Mtx toxins. This synergism has even been shown to overcome an insect's resistance to the other toxin. (Wirth 1997, Wirth 2005, Thiery 1998, Zhang 2006) The Cyt synergistic effect for Cry toxins is proposed to involve Cyt1A binding to domain II of Cry toxins in solution or on the membrane plane to promote formation of a Cry toxin pre-pore oligomer. Formation of this oligomer is independent of the Cyt oligomerization, binding or insertion. (Lailak 2013, Perez 2007, Lopez-Diaz 2013)

A number of pesticidal proteins unrelated to the Cry proteins are produced by some strains of B. thuringiensis and B. cereus during vegetative growth (Estruch et al. (1996) Proc Natl Acad Sci USA 93:5389-5394; Warren et al. (1994) WO 94/21795). These vegetative insecticidal proteins, or Vips, do not form parasporal crystal proteins and are apparently secreted from the cell. The Vips are presently excluded from the Cry protein nomenclature because they are not crystal-forming proteins. The term VIP is a misnomer in the sense that some B. thuringiensis Cry proteins are also produced during vegetative growth as well as during the stationary and sporulation phases, most notably Cry3Aa. The location of the Vip genes in the B. thuringiensis genome has been reported to reside on large plasmids that also encode cry genes (Mesrati et al. (2005) FEMS Microbiol. Lett. 244(2):353-8). A web-site for the nomenclature of Bt toxins can be found on the world wide web at lifesci.sussex.ac.uk with the path “/home/Neil_Crickmore/Bt/” and at: “btnomenclature.info/”. See also, Schnepf et al. (1998) Microbiol. Mol. Biol. Rev. 62(3):775-806. Such references are herein incorporated by reference.

To date four categories of Vips have been identified. Some Vip genes form binary two-component protein complexes; an “A” component is usually the “active” portion, and a “B” component is usually the “binding” portion. (Pfam pfam.xfam.org/family/PF03495). The Vip1 and Vip4 proteins generally contain binary toxin B protein domains. Vip2 proteins generally contain binary toxin A protein domains.

The Vip1 and Vip2 proteins are the two components of a binary toxin that exhibits toxicity to coleopterans. Vip1Aa1 and Vip2Aa1 are very active against corn rootworms, particularly Diabrotica virgifera and Diabrotica longicornis (Han et al. (1999) Nat. Struct. Biol. 6:932-936; Warren GW (1997) “Vegetative insecticidal proteins: novel proteins for control of corn pests” In: Carozzi NB, Koziel M (eds) Advances in insect control, the role of transgenic plants; Taylor & Francis Ltd, London, pp 109-21). The membrane-binding 95 kDa Vip1 multimer provides a pathway for the 52 kDa Vip2 ADP-ribosylase to enter the cytoplasm of target western corn rootworm cells (Warren (1997) supra). The NAD-dependent ADP-ribosyltransferase Vip2 likely modifies monomeric actin at Arg177 to block polymerization, leading to loss of the actin cytoskeleton and eventual cell death due to the rapid subunit ex-change within actin filaments in vivo (Carlier M. F. (1990) Adv. Biophys. 26:51-73).

Like Cry toxins, activated Vip3A toxins are pore-forming proteins capable of making stable ion channels in the membrane (Lee et al. (2003) Appl. Environ. Microbiol. 69:4648-4657). Vip3 proteins are active against several major lepidopteran pests (Rang et al. (2005) Appl. Environ. Microbiol. 71(10):6276-6281; Bhalla et al. (2005) FEMS Microbiol. Lett. 243:467-472; Estruch et al. (1998) WO 9844137; Estruch et al. (1996) Proc Natl Acad Sci USA 93:5389-5394; Selvapandiyan et al. (2001) Appl. Environ Microbiol. 67:5855-5858; Yu et al. (1997) Appl. Environ Microbiol. 63:532-536). Vip3A is active against Agrotis ipsilon, Spodoptera frupperda, Spodoptera exigua, Heliothis virescens, and Helicoverpa zea (Warren et al. (1996) WO 96/10083; Estruch et al. (1996) Proc Natl Acad Sci USA 93:5389-5394). Like Cry toxins, Vip3A proteins must be activated by proteases prior to recognition at the surface of the midgut epithelium of specific membrane proteins different from those recognized by Cry toxins.

The MTX family of toxin proteins is characterized by the presence of a conserved domain, ETX_MTX2 (pfam 03318). Members of this family share sequence homology with the mosquitocidal toxins Mtx2 and Mtx3 from Bacillus sphaericus, as well as with the epsilon toxin ETX from Clostridium perfringens (Cole et al. (2004) Nat. Struct. Mol. Biol. 11: 797-8; Thanabalu et al. (1996) Gene 170:85-9). The MTX-like proteins are structurally distinct from the three-domain Cry toxins, as they have an elongated and predominately β-sheet-based structure. However, similar to the three-domain toxins, the MTX-like proteins are thought to form pores in the membranes of target cells (Adang et al. (2014) supra). Unlike the three-domain Cry proteins, the MTX-like proteins are much smaller in length, ranging from 267 amino acids (Cry23) to 340 amino acids (Cry15A).

To date, only 15 proteins belonging to the family of MTX-like toxins have been assigned Cry names, making this a relatively small class compared to the three-domain Cry family (Crickmore et al. (2014) supra; Adang et al. (2014) supra). The members of the MTX-like toxin family include Cry15, Cry23, Cry33, Cry38, Cry45, Cry46, Cry51, Cry60A, Cry60B, and Cry64. This family exhibits a range of insecticidal activity, including activity against insect pests of the Lepidopteran and Coleopteran orders. Some members of this family may form binary partnerships with other proteins, which may or may not be required for insecticidal activity.

Cry15 is a 34 kDA protein that was identified in Bacillus thuringiensis serovar thompsoni HD542; it occurs naturally in a crystal together with an unrelated protein of approximately 40 kDa. The gene encoding Cry15 and its partner protein are arranged together in an operon. Cry15 alone has been shown to have activity against lepidopteran insect pests including Manduca sexta, Cydia pomonella, and Pieris rapae, with the presence of the 40 kDA protein having been shown to increase activity of Cry15 only against C. pomonella (Brown K. and Whiteley H. (1992) J. Bacteriol. 174:549-557; Naimov et al. (2008) Appl. Environ. Microbiol. 74:7145-7151). Further studies are needed to elucidate the function of the partner protein of Cry15. Similarly, Cry23 is a 29 kDA protein that has been shown to have activity against the coleopteran pests Tribolium castaneum and Popillia japonica together with its partner protein Cry37 (Donovan et al. (2000) U.S. Pat. No. 6,063,756).

New members of the MTX-like family are continuing to be identified. An ETX_MTX toxin gene was recently identified in the genome of Bacillus thuringiensis serovar tolworthi strain Na205-3. This strain was found to be toxic against the lepidpoteran pest Helicoverpa armigera, and it also contained homologs of Cry1, Cry11, Vip1, Vip2, and Vip3 (Palma et al. (2014) Genome Announc. 2(2): e00187-14. Published online Mar. 13, 2014 at doi: 10.1128/genomeA.00187-14; PMCID: PMC3953196). Because the MTX-like proteins have a unique domain structure relative to the three-domain Cry proteins, they are believed to possess a unique mode of action, thereby making them a valuable tool in insect control and the fight against insect resistance.

Bacterial cells produce large numbers of toxins with diverse specificity against host and non-host organisms. Large families of binary toxins have been identified in numerous bacterial families, including toxins that have activity against insect pests. (Poopathi and Abidha (2010) J. Physiol. Path. 1(3): 22-38). Lysinibacillus sphaericus (Ls), formerly Bacillus sphaericus, (Ahmed et al. (2007) Int. J Syst. Evol. Microbiol. 57:1117-1125) is well-known as an insect biocontrol strain. Ls produces several insecticidal proteins, including the highly potent binary complex BinA/BinB. This binary complex forms a parasporal crystal in Ls cells and has strong and specific activity against dipteran insects, specifically mosquitos. In some areas, insect resistance to existing Ls mosquitocidal strains has been reported. The discovery of new binary toxins with different target specificity or the ability to overcome insect resistance is of significant interest.

The Ls binary insecticidal protein complex contains two major polypeptides, a 42 kDa polypeptide and a 51 kDa polypepdide, designated BinA and BinB, respectively (Ahmed et al. (2007) supra). The two polypeptides act synergistically to confer toxicity to their targets. Mode of action involves binding of the proteins to receptors in the larval midgut. In some cases, the proteins are modified by protease digestion in the larval gut to produce activated forms. The BinB component is thought to be involved in binding, while the BinA component confers toxicity (Nielsen-LeRoux et al. (2001) Appl. Environ. Microbiol. 67(11):5049-5054). When cloned and expressed separately, the BinA component is toxic to mosquito larvae, while the BinB component is not. However, co-administration of the proteins markedly increases toxicity (Nielsen-LeRoux et al. (2001) supra).

A small number of Bin protein homologs have been described from bacterial sources. Priest et al. (1997) Appl. Environ. Microbiol. 63(4):1195-1198 describe a hybridization effort to identify new Ls strains, although most of the genes they identified encoded proteins identical to the known BinA/BinB proteins. The BinA protein contains a defined conserved domain known as the Toxin 10 superfamily domain. This toxin domain was originally defined by its presence in BinA and BinB. The two proteins both have the domain, although the sequence similarity between BinA and BinB is limited in this region (<40%). The Cry49Aa protein, which also has insecticidal activity, also has this domain (described below).

The Cry48Aa/Cry49Aa binary toxin of Ls has the ability to kill Culex quinquefasciatus mosquito larvae. These proteins are in a protein structural class that has some similarity to the Cry protein complex of Bacillus thuringiensis (Bt), a well-known insecticidal protein family. The Cry34/Cry35 binary toxin of Bt is also known to kill insects, including Western corn rootworm, a significant pest of corn. Cry34, of which several variants have been identified, is a small (14 kDa) polypeptide, while Cry35 (also encoded by several variants) is a 44 kDa polypeptide. These proteins have some sequence homology with the BinA/BinB protein group and are thought to be evolutionarily related (Ellis et al. (2002) Appl. Environ. Microbiol. 68(3):1137-1145).

Provided herein are pesticidal proteins from these classes of toxins. The pesticidal proteins are classified by their structure, homology to known toxins and/or their pesticidal specificity.

ii. Variants and Fragments of Pesticidal Proteins and Polynucleotides Encoding the Same

Pesticidal proteins or polypeptides of the invention include those set forth in SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398 and fragments and variants thereof. By “pesticidal toxin” or “pesticidal protein” or “pesticidal polypeptide” is intended a toxin or protein or polypeptide that has activity against one or more pests, including, insects, fungi, nematodes, and the like such that the pest is killed or controlled.

An “isolated” or “purified” polypeptide or protein, or biologically active portion thereof, is substantially or essentially free from components that normally accompany or interact with the polypeptide or protein as found in its naturally occurring environment. Thus, an isolated or purified polypeptide or protein is substantially free of other cellular material, or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized. A protein that is substantially free of cellular material includes preparations of protein having less than about 30%, 20%, 10%, 5%, or 1% (by dry weight) of contaminating protein. When the protein of the invention or biologically active portion thereof is recombinantly produced, optimally culture medium represents less than about 30%, 20%, 10%, 5%, or 1% (by dry weight) of chemical precursors or non-protein-of-interest chemicals.

The term “fragment” refers to a portion of a polypeptide sequence of the invention. “Fragments” or “biologically active portions” include polypeptides comprising a sufficient number of contiguous amino acid residues to retain the biological activity, i.e., have pesticidal activity. Fragments of the pesticidal proteins include those that are shorter than the full-length sequences, either due to the use of an alternate downstream start site, or due to processing that produces a shorter protein having pesticidal activity. Processing may occur in the organism the protein is expressed in, or in the pest after ingestion of the protein. Examples of fragments of the proteins can be found in Table 1. A biologically active portion of a pesticidal protein can be a polypeptide that is, for example, 10, 25, 50, 100, 150, 200, 250 or more amino acids in length of any one of SEQ ID NO:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398. Such biologically active portions can be prepared by recombinant techniques and evaluated for pesticidal activity. As used here, a fragment comprises at least 8 contiguous amino acids of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398.

Bacterial genes, including those encoding the pesticidal proteins disclosed herein, quite often possess multiple methionine initiation codons in proximity to the start of the open reading frame. Often, translation initiation at one or more of these start codons will lead to generation of a functional protein. These start codons can include ATG codons. However, bacteria such as Bacillus sp. also recognize the codon GTG as a start codon, and proteins that initiate translation at GTG codons contain a methionine at the first amino acid. On rare occasions, translation in bacterial systems can initiate at a TTG codon, though in this event the TTG encodes a methionine. Furthermore, it is not often determined a priori which of these codons are used naturally in the bacterium. Thus, it is understood that use of one of the alternate methionine codons may also lead to generation of pesticidal proteins. These pesticidal proteins are encompassed in the present invention and may be used in the methods disclosed herein. It will be understood that, when expressed in plants, it will be necessary to alter the alternate start codon to ATG for proper translation.

In various embodiments the pesticidal proteins provided herein include amino acid sequences deduced from the full-length nucleotide sequences and amino acid sequences that are shorter than the full-length sequences due to the use of an alternate downstream start site. Thus, the nucleotide sequence of the invention and/or vectors, host cells, and plants comprising the nucleotide sequence of the invention (and methods of making and using the nucleotide sequence of the invention) may comprise a nucleotide sequence encoding an alternate start site.

It is recognized that modifications may be made to the pesticidal polypeptides provided herein creating variant proteins. Changes designed by man may be introduced through the application of site-directed mutagenesis techniques. Alternatively, native, as yet-unknown or as yet unidentified polynucleotides and/or polypeptides structurally and/or functionally-related to the sequences disclosed herein may also be identified that fall within the scope of the present invention. Conservative amino acid substitutions may be made in nonconserved regions that do not alter the function of the pesticidal proteins. Alternatively, modifications may be made that improve the activity of the toxin. Modification of Cry toxins by domain III swapping has resulted in some cases in hybrid toxins with improved toxicities against certain insect species. Thus, domain III swapping could be an effective strategy to improve toxicity of Cry toxins or to create novel hybrid toxins with toxicity against pests that show no susceptibility to the parental Cry toxins. Site-directed mutagenesis of domain II loop sequences may result in new toxins with increased insecticidal activity. Domain II loop regions are key binding regions of initial Cry toxins that are suitable targets for the mutagenesis and selection of Cry toxins with improved insecticidal properties. Domain I of the Cry toxin may be modified to introduce protease cleavage sites to improve activity against certain pests. Strategies for shuffling the three different domains among large numbers of cry genes and high throughput bioassay screening methods may provide novel Cry toxins with improved or novel toxicities.

As indicated, fragments and variants of the polypeptides disclosed herein will retain pesticidal activity. Pesticidal activity comprises the ability of the composition to achieve an observable effect diminishing the occurrence or an activity of the target pest, including for example, bringing about death of at least one pest, or a noticeable reduction in pest growth, feeding, or normal physiological development. Such decreases in numbers, pest growth, feeding or normal development can comprise any statistically significant decrease, including, for example a decrease of about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 85%, 90%, 95% or greater. It is recognized that the pesticidal activity may be different or improved relative to the activity of the native protein, or it may be unchanged, so long as pesticidal activity is retained. Methods for measuring pesticidal activity are well known in the art. See, for example, Czapla and Lang (1990) J. Econ. Entomol. 83:2480-2485; Andrews et al. (1988) Biochem. J. 252:199-206; Marrone et al. (1985) J. of Economic Entomology 78:290-293; and U.S. Pat. No. 5,743,477, all of which are herein incorporated by reference in their entirety.

Polypeptide variants of this disclosure include polypeptides having an amino acid sequence that is at least about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98% or about 99% identical to the amino acid sequence of any of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398 and retain pesticidal activity. Note, Table 1 provides non-limiting examples of variant polypeptides (and polynucleotide encoding the same) for each of SEQ ID NOS: 1-398. A biologically active variant of a pesticidal polypeptide of the invention may differ by as few as about 1-15 amino acid residues, as few as about 1-10, such as about 6-10, as few as 5, as few as 4, as few as 3, as few as 2, or as few as 1 amino acid residue. In specific embodiments, the polypeptides can comprise an N′-terminal or a C′-terminal truncation, which can comprise at least a deletion of 10, 15, 20, 25, 30, 35, 40, 45, 50 amino acids or more from either the N′ or C′ terminal end of the polypeptide.

Table 2 provides protein domains found in SEQ ID NOs: 1-398 based on PFAM data. Both the domain description and the positions within a given SEQ ID NO are provided in Table 2. In specific embodiments, the active variant comprising any one of SEQ ID NOs: 1-398 can comprise at least 70%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any one of SEQ ID NOs: 1-398 and further comprises at least one of the conserved domain set forth in Table 2. For example, in one embodiment, the active variant will comprise at least 70%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO:1, and further comprises the native amino acids at positions 54-304.

TABLE 2 Summary of PFAM domains in each of SEQ ID NOs: 1-398 Domain Modification Domain Position APG ID Seq ID Type PFAM Domain Description Start Stop APG00056 Seq ID 1 PF03945 Endotoxin N 54 304 PF00555 Endotoxin M 311 500 PF03944 Endotoxin C 510 646 APG00056 Seq ID 2 3′ Truncation PF03945 Endotoxin N 54 304 modified PF00555 Endotoxin M 311 500 PF03944 Endotoxin C 510 645 APG00058 Seq ID 3 PF03945 Endotoxin N 68 320 PF00555 Endotoxin M 327 513 PF03944 Endotoxin C 523 658 APG00058 Seq ID 4 3′ Truncation PF03945 Endotoxin N 68 320 modified PF00555 Endotoxin M 327 513 PF03944 Endotoxin C 523 657 APG00064 Seq ID 5 PF03318 ETX MTX2 94 329 APG00064 Seq ID 6 Signal peptide PF03318 ETX MTX2 67 302 modified removed APG00067 Seq ID 7 PF03318 ETX MTX2 29 252 APG00071 Seq ID 8 PF12495 Vip3A N 12 188 APG00071 Seq ID 9 Alternate start PF12495 Vip3A N 9 185 modified APG00073 Seq ID 10 PF03945 Endotoxin N 5 244 PF03944 Endotoxin C 478 612 APG00073 Seq ID 11 Alternate start PF03945 Endotoxin N 5 244 modified and 3′ PF03944 Endotoxin C 478 611 Truncation APG00073 Seq ID 12 Alternate start PF03945 Endotoxin N 5 244 modified PF03944 Endotoxin C 478 612 APG00073 Seq ID 13 3′ Truncation PF03945 Endotoxin N 5 244 modified PF03944 Endotoxin C 478 611 APG00074 Seq ID 14 PF12495 Vip3A N 16 191 APG00084 Seq ID 15 PF03945 Endotoxin N 76 300 PF00555 Endotoxin M 305 529 PF03944 Endotoxin C 539 673 APG00084 Seq ID 16 3′ Truncation PF03945 Endotoxin N 76 300 modified PF00555 Endotoxin M 305 529 PF03944 Endotoxin C 539 672 APG00105 Seq ID 17 PF03945 Endotoxin N 95 330 PF00555 Endotoxin M 335 542 PF03944 Endotoxin C 552 694 APG00105 Seq ID 18 Alternate start PF03945 Endotoxin N 64 299 modified PF00555 Endotoxin M 304 511 PF03944 Endotoxin C 521 663 APG00105 Seq ID 19 Alternate start PF03945 Endotoxin N 64 299 modified and 3′ PF00555 Endotoxin M 304 511 Truncation PF03944 Endotoxin C 521 662 APG00105 Seq ID 20 3′ Truncation PF03945 Endotoxin N 95 330 modified PF00555 Endotoxin M 335 542 PF03944 Endotoxin C 552 693 APG00107 Seq ID 21 PF03318 ETX MTX2 29 250 APG00108 Seq ID 22 PF01338 Bac thur toxin 72 265 APG00112 Seq ID 23 PF03318 ETX MTX2 28 267 APG00113 Seq ID 24 PF03945 Endotoxin N 56 302 PF00030 Crystall 653 732 PF00030 Crystall 733 814 PF14200 RicinB lectin 2 855 962 APG00116 Seq ID 25 PF03945 Endotoxin N 72 294 PF00555 Endotoxin M 299 502 PF03944 Endotoxin C 512 645 APG00116 Seq ID 26 3′ Truncation PF03945 Endotoxin N 72 294 modified PF00555 Endotoxin M 299 502 PF03944 Endotoxin C 512 644 APG00117 Seq ID 27 PF01338 Bac thur toxin 378 572 APG00118 Seq ID 28 PF05431 Toxin 10 237 434 APG00121 Seq ID 29 PF01338 Bac thur toxin 52 272 APG00131 Seq ID 30 PF12495 Vip3A N 10 187 APG00132 Seq ID 31 PF05431 Toxin 10 209 402 APG00134 Seq ID 32 PF01338 Bac thur toxin 2 191 APG00137 Seq ID 33 PF03318 ETX MTX2 21 256 APG00141 Seq ID 34 PF05431 Toxin 10 183 383 APG00150 Seq ID 35 PF12495 Vip3A N 12 188 APG00152 Seq ID 36 PF03945 Endotoxin N 70 293 PF00555 Endotoxin M 298 522 PF03944 Endotoxin C 532 683 APG00152 Seq ID 37 3′ Truncation PF03945 Endotoxin N 70 293 modified PF00555 Endotoxin M 298 522 PF03944 Endotoxin C 532 682 APG00153 Seq ID 38 PF03945 Endotoxin N 101 323 PF00555 Endotoxin M 328 531 PF03944 Endotoxin C 541 670 APG00153 Seq ID 39 3′ Truncation PF03945 Endotoxin N 101 323 modified PF00555 Endotoxin M 328 531 PF03944 Endotoxin C 541 669 APG00155 Seq ID 40 PF03318 ETX MTX2 25 250 APG00164 Seq ID 41 PF03945 Endotoxin N 56 279 PF00555 Endotoxin M 284 502 PF03944 Endotoxin C 512 649 APG00164 Seq ID 42 3′ Truncation PF03945 Endotoxin N 56 279 modified PF00555 Endotoxin M 284 502 PF03944 Endotoxin C 512 648 APG00166 Seq ID 43 PF12495 Vip3A N 16 188 APG00168 Seq ID 44 PF01338 Bac thur toxin 35 257 APG00172 Seq ID 45 PF03318 ETX MTX2 36 238 APG00173 Seq ID 46 PF12495 Vip3A N 16 188 APG00173 Seq ID 47 Alternate start PF12495 Vip3A N 14 186 modified APG00175 Seq ID 48 PF12495 Vip3A N 16 188 PF02018 CBM 4 9 549 663 APG00175 Seq ID 49 Alternate start PF12495 Vip3A N 14 186 modified PF02018 CBM 4 9 547 661 APG00176 Seq ID 50 PF03945 Endotoxin N 58 280 PF00555 Endotoxin M 285 490 PF03944 Endotoxin C 500 637 APG00176 Seq ID 51 3′ Truncation PF03945 Endotoxin N 58 280 modified PF00555 Endotoxin M 285 490 PF03944 Endotoxin C 500 636 APG00177 Seq ID 52 PF01338 Bac thur toxin 27 250 APG00177 Seq ID 53 3′ Truncation PF01338 Bac thur toxin 27 224 modified APG00178 Seq ID 54 PF05431 Toxin 10 187 379 APG00180 Seq ID 55 PF05791 Bacillus HBL 60 201 APG00186 Seq ID 56 PF01338 Bac thur toxin 378 572 APG00188 Seq ID 57 PF03945 Endotoxin N 62 323 PF00555 Endotoxin M 331 512 PF03944 Endotoxin C 527 669 APG00188 Seq ID 58 3′ Truncation PF03945 Endotoxin N 62 323 modified PF00555 Endotoxin M 331 512 PF03944 Endotoxin C 527 668 APG00189 Seq ID 59 PF03945 Endotoxin N 38 274 PF00555 Endotoxin M 279 470 PF03944 Endotoxin C 480 617 APG00189 Seq ID 60 Alternate start PF03945 Endotoxin N 29 265 modified PF00555 Endotoxin M 270 461 PF03944 Endotoxin C 471 608 APG00189 Seq ID 61 3′ Truncation PF03945 Endotoxin N 38 274 modified PF00555 Endotoxin M 279 470 PF03944 Endotoxin C 480 616 APG00190 Seq ID 62 PF03945 Endotoxin N 68 291 PF00555 Endotoxin M 296 522 PF03944 Endotoxin C 532 676 APG00190 Seq ID 63 3′ Truncation PF03945 Endotoxin N 68 291 modified PF00555 Endotoxin M 296 522 PF03944 Endotoxin C 532 675 APG00192 Seq ID 64 PF05431 Toxin 10 159 358 APG00194 Seq ID 65 PF01338 Bac thur toxin 27 223 APG00196 Seq ID 66 PF03945 Endotoxin N 58 302 PF00030 Crystall 653 732 PF00030 Crystall 733 814 PF14200 RicinB lectin 2 855 962 APG00198 Seq ID 67 PF01338 Bac thur toxin 2 190 APG00200 Seq ID 68 PF01338 Bac thur toxin 282 482 APG00203 Seq ID 69 PF05431 Toxin 10 189 382 APG00207 Seq ID 70 PF12495 Vip3A N 16 193 APG00209 Seq ID 71 PF05791 Bacillus HBL 40 226 APG00211 Seq ID 73 PF14200 RicinB lectin 2 3 102 PF05431 Toxin 10 156 353 APG00211 Seq ID 74 Alternate start PF14200 RicinB lectin 2 2 98 modified PF05431 Toxin 10 152 349 APG00212 Seq ID 75 PF05431 Toxin 10 216 409 APG00213 Seq ID 76 PF05431 Toxin 10 188 385 APG00214 Seq ID 77 PF05791 Bacillus HBL 48 231 APG00215 Seq ID 78 PF14200 RicinB lectin 2 2 96 PF05431 Toxin 10 148 343 APG00216 Seq ID 79 PF00652 Ricin B lectin 38 165 PF05431 Toxin 10 175 287 APG00219 Seq ID 80 PF03945 Endotoxin N 116 337 APG00221 Seq ID 81 PF05431 Toxin 10 192 385 APG00223 Seq ID 82 PF05431 Toxin 10 235 432 APG00227 Seq ID 83 PF03318 ETX MTX2 150 383 APG00229 Seq ID 84 PF05431 Toxin 10 210 403 APG00230 Seq ID 85 PF00652 Ricin B lectin 36 167 PF05431 Toxin 10 177 373 APG00231 Seq ID 86 PF14200 RicinB lectin 2 47 150 PF05431 Toxin 10 156 353 APG00232 Seq ID 87 PF03945 Endotoxin N 110 310 PF14200 RicinB lectin 2 560 662 APG00233 Seq ID 88 PF03945 Endotoxin N 86 292 PF14200 RicinB lectin 2 404 513 APG00235 Seq ID 89 PF03945 Endotoxin N 128 345 PF14200 RicinB lectin 2 462 565 APG00237 Seq ID 90 PF05431 Toxin 10 200 393 APG00239 Seq ID 91 PF03945 Endotoxin N 152 374 PF01473 CW binding 1 432 450 PF01473 CW binding 1 453 472 APG00241 Seq ID 92 PF05791 Bacillus HBL 44 225 APG00242 Seq ID 93 PF05431 Toxin 10 235 431 APG00243 Seq ID 94 PF05431 Toxin 10 188 381 APG00248 Seq ID 95 PF05431 Toxin 10 224 416 APG00249 Seq ID 96 PF05431 Toxin 10 219 413 APG00251 Seq ID 97 PF03945 Endotoxin N 73 279 PF14200 RicinB lectin 2 391 500 APG00255 Seq ID 98 PF03945 Endotoxin N 105 337 PF01473 CW binding 1 384 402 PF14200 RicinB lectin 2 484 587 APG00258 Seq ID 99 PF05791 Bacillus HBL 80 219 APG00261 Seq ID 100 PF05431 Toxin 10 196 393 APG00262 Seq ID 101 PF05431 Toxin 10 235 429 APG00264 Seq ID 103 PF05791 Bacillus HBL 49 229 APG00266 Seq ID 105 PF03945 Endotoxin N 165 358 PF01473 CW binding 1 453 472 PF01473 CW binding 1 474 489 APG00267 Seq ID 106 PF03945 Endotoxin N 122 321 APG00271 Seq ID 108 PF05791 Bacillus HBL 32 209 APG00273 Seq ID 109 PF12495 Vip3A N 16 188 PF02018 CBM 4 9 545 658 APG00274 Seq ID 110 PF05791 Bacillus HBL 56 238 APG00275 Seq ID 111 PF03945 Endotoxin N 5 129 APG00278 Seq ID 112 PF12495 Vip3A N 16 188 APG00278 Seq ID 113 Alternate start PF12495 Vip3A N 14 186 modified APG00279 Seq ID 114 PF05791 Bacillus HBL 128 239 APG00282 Seq ID 116 PF05431 Toxin 10 167 362 APG00284 Seq ID 117 PF14200 RicinB lectin 2 47 150 PF05431 Toxin 10 156 353 APG00285 Seq ID 118 PF03945 Endotoxin N 108 338 PF01473 CW binding 1 407 421 PF01473 CW binding 1 456 470 PF01473 CW binding 1 573 589 APG00286 Seq ID 119 PF05791 Bacillus HBL 37 217 APG00286 Seq ID 120 3′ Truncation PF05791 Bacillus HBL 37 217 modified APG00287 Seq ID 121 PF14200 RicinB lectin 2 53 150 PF05431 Toxin 10 156 353 APG00288 Seq ID 122 PF03945 Endotoxin N 116 318 PF01473 CW binding 1 410 429 PF01473 CW binding 1 431 445 APG00290 Seq ID 124 PF01338 Bac thur toxin 55 281 APG00292 Seq ID 126 PF03945 Endotoxin N 95 304 PF05588 Botulinum 401 501 HA-17 APG00294 Seq ID 127 PF05791 Bacillus HBL 46 229 APG00295 Seq ID 128 PF05431 Toxin 10 224 420 APG00297 Seq ID 129 PF05791 Bacillus HBL 63 243 APG00298 Seq ID 130 PF05791 Bacillus HBL 38 218 APG00301 Seq ID 131 PF05431 Toxin 10 189 382 APG00302 Seq ID 132 PF03945 Endotoxin N 103 312 PF14200 RicinB lectin 2 424 519 APG00305 Seq ID 133 PF05791 Bacillus HBL 47 233 APG00307 Seq ID 134 PF05791 Bacillus HBL 43 228 APG00308 Seq ID 135 PF05431 Toxin 10 203 397 APG00314 Seq ID 136 PF05431 Toxin 10 186 380 APG00315 Seq ID 137 PF05431 Toxin 10 188 380 APG00320 Seq ID 140 PF05431 Toxin 10 252 446 APG00340 Seq ID 141 PF14200 RicinB lectin 2 69 170 PF05431 Toxin 10 176 374 APG00342 Seq ID 142 PF05791 Bacillus HBL 53 205 APG00349 Seq ID 143 PF14200 RicinB lectin 2 1 97 PF05431 Toxin 10 154 347 APG00353 Seq ID 144 PF05431 Toxin 10 189 382 APG00355 Seq ID 145 PF03945 Endotoxin N 56 270 APG00355 Seq ID 146 3′ Truncation PF03945 Endotoxin N 55 270 modified APG00356 Seq ID 147 PF14200 RicinB lectin 2 53 150 PF05431 Toxin 10 156 353 APG00368 Seq ID 148 PF14200 RicinB lectin 2 3 102 PF05431 Toxin 10 156 354 APG00369 Seq ID 149 PF05431 Toxin 10 251 445 APG00372 Seq ID 150 PF03945 Endotoxin N 119 304 PF01473 CW binding 1 420 439 PF01473 CW binding 1 441 455 APG00373 Seq ID 151 PF00652 Ricin B lectin 38 165 PF05431 Toxin 10 176 372 APG00375 Seq ID 152 PF03945 Endotoxin N 117 335 PF01473 CW binding 1 458 475 APG00376 Seq ID 153 PF05431 Toxin 10 192 384 APG00377 Seq ID 154 PF14200 RicinB lectin 2 54 150 PF05431 Toxin 10 156 353 APG00379 Seq ID 155 PF14200 RicinB lectin 2 47 148 PF05431 Toxin 10 156 353 APG00385 Seq ID 157 PF05431 Toxin 10 203 396 APG00386 Seq ID 158 PF14200 RicinB lectin 2 42 143 PF05431 Toxin 10 149 343 APG00388 Seq ID 159 PF01338 Bac thur toxin 9 233 APG00391 Seq ID 160 PF03945 Endotoxin N 110 306 APG00392 Seq ID 161 PF05431 Toxin 10 203 396 APG00395 Seq ID 162 PF03945 Endotoxin N 111 332 PF01473 CW binding 1 433 449 PF01473 CW binding 1 461 477 APG00396 Seq ID 163 PF03945 Endotoxin N 115 310 PF01473 CW binding 1 401 418 PF01473 CW binding 1 420 439 PF01473 CW binding 1 523 538 APG00398 Seq ID 165 PF05431 Toxin 10 278 475 APG00399 Seq ID 166 PF00041 fn3 430 523 PF14200 RicinB lectin 2 1068 1192 PF05431 Toxin 10 1364 1558 APG00401 Seq ID 168 PF03945 Endotoxin N 104 336 PF01473 CW binding 1 383 401 PF14200 RicinB lectin 2 483 586 APG00404 Seq ID 169 PF03945 Endotoxin N 48 264 PF01473 CW binding 1 346 365 PF01473 CW binding 1 388 406 PF01473 CW binding 1 460 476 APG00407 Seq ID 170 PF05431 Toxin 10 208 401 APG00408 Seq ID 171 PF03318 ETX MTX2 29 252 APG00409 Seq ID 172 PF01338 Bac thur toxin 63 261 APG00412 Seq ID 173 PF05431 Toxin 10 197 390 APG00413 Seq ID 174 PF00652 Ricin B lectin 36 162 PF05431 Toxin 10 172 371 APG00415 Seq ID 175 PF05431 Toxin 10 154 347 APG00419 Seq ID 176 PF05431 Toxin 10 203 396 APG00422 Seq ID 177 PF01338 Bac thur toxin 21 244 APG00427 Seq ID 178 PF03318 ETX MTX2 83 314 APG00454 Seq ID 179 PF05431 Toxin 10 235 431 APG00456 Seq ID 180 PF01338 Bac thur toxin 1 200 APG00459 Seq ID 181 PF05431 Toxin 10 211 404 APG00461 Seq ID 182 PF05431 Toxin 10 258 455 APG00462 Seq ID 183 PF01338 Bac thur toxin 7 228 APG00467 Seq ID 184 PF03945 Endotoxin N 75 287 APG00499 Seq ID 185 PF00652 Ricin B lectin 24 153 PF05431 Toxin 10 164 337 APG00500 Seq ID 186 PF03945 Endotoxin N 57 301 APG00517 Seq ID 187 PF03318 ETX MTX2 72 305 APG00521 Seq ID 188 PF05431 Toxin 10 194 387 APG00532 Seq ID 189 PF14200 RicinB lectin 2 66 156 PF05431 Toxin 10 163 358 APG00550 Seq ID 190 PF00652 Ricin B lectin 9 131 PF05431 Toxin 10 148 337 APG00559 Seq ID 191 PF03318 ETX MTX2 6 224 APG00571 Seq ID 192 PF14200 RicinB lectin 2 4 102 PF05431 Toxin 10 156 350 APG00590 Seq ID 193 PF03318 ETX MTX2 74 294 APG00591 Seq ID 194 PF05431 Toxin 10 155 349 APG00592 Seq ID 195 PF05431 Toxin 10 216 409 APG00596 Seq ID 196 PF00652 Ricin B lectin 37 178 PF05431 Toxin 10 179 379 APG00600 Seq ID 197 PF05431 Toxin 10 207 399 APG00603 Seq ID 198 PF03945 Endotoxin N 69 279 APG00619 Seq ID 199 PF05431 Toxin 10 207 399 APG00635 Seq ID 201 PF03318 ETX MTX2 126 252 APG00642 Seq ID 202 PF00652 Ricin B lectin 9 142 PF05431 Toxin 10 153 347 APG00642 Seq ID 203 3′ Truncation PF00652 Ricin B lectin 9 142 modified APG00647 Seq ID 204 PF01338 Bac thur toxin 50 275 APG00652 Seq ID 205 PF05791 Bacillus HBL 40 232 APG00698 Seq ID 206 PF01338 Bac thur toxin 52 280 APG00700 Seq ID 207 PF05431 Toxin 10 192 383 APG00723 Seq ID 209 PF03945 Endotoxin N 171 379 APG00727 Seq ID 210 PF05431 Toxin 10 187 379 APG00731 Seq ID 211 PF14200 RicinB lectin 2 47 150 PF05431 Toxin 10 156 353 APG00732 Seq ID 212 PF00652 Ricin B lectin 24 153 PF05431 Toxin 10 163 358 PF00388 PI-PLC-X 406 548 PF14200 RicinB lectin 2 837 939 APG00741 Seq ID 213 PF00652 Ricin B lectin 39 160 PF05431 Toxin 10 170 370 APG00748 Seq ID 214 PF14200 RicinB lectin 2 42 143 PF05431 Toxin 10 149 346 APG00757 Seq ID 215 PF00652 Ricin B lectin 63 184 PF05431 Toxin 10 194 394 APG00767 Seq ID 216 PF05431 Toxin 10 203 411 APG00768 Seq ID 217 PF03318 ETX MTX2 99 335 APG00797 Seq ID 218 PF14200 RicinB lectin 2 43 147 PF05431 Toxin 10 153 347 APG00798 Seq ID 219 PF05431 Toxin 10 216 408 APG00830 Seq ID 220 PF05791 Bacillus HBL 47 238 APG00844 Seq ID 221 PF05431 Toxin 10 188 381 APG00851 Seq ID 222 PF03318 ETX MTX2 82 313 APG00862 Seq ID 223 PF14200 RicinB lectin 2 2 57 PF05431 Toxin 10 63 259 APG00898 Seq ID 224 PF03318 ETX MTX2 30 254 APG00907 Seq ID 225 PF03318 ETX MTX2 28 266 APG00913 Seq ID 226 PF05431 Toxin 10 252 449 APG00925 Seq ID 227 PF05431 Toxin 10 242 437 APG00074 Seq ID 228 Alternate start PF12495 Vip3A N 12 187 modified APG00084 Seq ID 229 Alternate start PF03945 Endotoxin N 66 290 modified PF00555 Endotoxin M 295 519 PF03944 Endotoxin C 529 663 APG00108 Seq ID 230 Alternate start PF01338 Bac thur toxin 1 191 modified APG00113 Seq ID 231 Alternate start PF03945 Endotoxin N 53 299 modified PF00030 Crystall 650 729 PF00030 Crystall 730 811 PF14200 RicinB lectin 2 852 959 APG00113 Seq ID 232 Alternate start PF03945 Endotoxin N 25 271 modified PF00030 Crystall 622 701 PF00030 Crystall 702 783 PF14200 RicinB lectin 2 824 931 APG00116 Seq ID 233 Alternate start PF03945 Endotoxin N 69 291 modified PF00555 Endotoxin M 296 499 PF03944 Endotoxin C 509 642 APG00116 Seq ID 234 Alternate start PF03945 Endotoxin N 69 291 modified and 3′ PF00555 Endotoxin M 296 499 Truncation PF03944 Endotoxin C 509 641 APG00117 Seq ID 235 Alternate start PF01338 Bac thur toxin 378 572 modified APG00117 Seq ID 236 Alternate start PF01338 Bac thur toxin 1 192 modified APG00118 Seq ID 237 Alternate start PF05431 Toxin 10 195 392 modified APG00121 Seq ID 238 Alternate start PF01338 Bac thur toxin 18 238 modified APG00132 Seq ID 239 Signal peptide PF05431 Toxin 10 180 373 modified removed APG00134 Seq ID 240 Alternate start PF01338 Bac thur toxin 1 191 modified APG00137 Seq ID 241 Alternate start PF03318 ETX MTX2 14 249 modified APG00141 Seq ID 242 Signal peptide PF05431 Toxin 10 149 349 modified removed APG00150 Seq ID 243 Alternate start PF12495 Vip3A N 9 185 modified APG00153 Seq ID 244 Alternate start PF03945 Endotoxin N 65 287 modified PF00555 Endotoxin M 292 495 PF03944 Endotoxin C 505 634 APG00153 Seq ID 245 Alternate start PF03945 Endotoxin N 65 287 modified and 3′ PF00555 Endotoxin M 292 495 Truncation PF03944 Endotoxin C 505 633 APG00166 Seq ID 246 Alternate start PF12495 Vip3A N 14 186 modified APG00168 Seq ID 247 Alternate start PF01338 Bac thur toxin 22 244 modified APG00177 Seq ID 248 Alternate start PF01338 Bac thur toxin 16 239 modified APG00186 Seq ID 249 Alternate start PF01338 Bac thur toxin 378 572 modified APG00186 Seq ID 250 Alternate start PF01338 Bac thur toxin 1 192 modified APG00194 Seq ID 251 Alternate start PF01338 Bac thur toxin 10 206 modified APG00196 Seq ID 252 Alternate start PF03945 Endotoxin N 55 299 modified PF00030 Crystall 650 729 PF00030 Crystall 730 811 PF14200 RicinB lectin 2 852 959 APG00198 Seq ID 253 Alternate start PF01338 Bac thur toxin 1 190 modified APG00200 Seq ID 254 Alternate start PF01338 Bac thur toxin 282 482 modified APG00200 Seq ID 255 Alternate start PF01338 Bac thur toxin 1 192 modified APG00203 Seq ID 256 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00209 Seq ID 257 Signal peptide PF05791 Bacillus HBL 10 196 modified removed APG00210 Seq ID 258 Alternate start no PFAM modified domains APG00212 Seq ID 259 Alternate start PF05431 Toxin 10 207 400 modified APG00212 Seq ID 260 Signal peptide PF05431 Toxin 10 178 371 modified removed APG00213 Seq ID 261 Signal peptide PF05431 Toxin 10 161 358 modified removed APG00214 Seq ID 262 Alternate start PF05791 Bacillus HBL 48 231 modified APG00219 Seq ID 263 Signal peptide PF03945 Endotoxin N 78 299 modified removed APG00221 Seq ID 264 Signal peptide PF05431 Toxin 10 165 358 modified removed APG00223 Seq ID 265 Alternate start PF05431 Toxin 10 193 390 modified APG00223 Seq ID 266 Signal peptide PF05431 Toxin 10 166 363 modified removed APG00227 Seq ID 267 Alternate start PF03318 ETX MTX2 122 355 modified APG00227 Seq ID 268 Signal peptide PF03318 ETX MTX2 93 326 modified removed APG00229 Seq ID 269 Alternate start PF05431 Toxin 10 201 394 modified APG00229 Seq ID 270 Signal peptide PF05431 Toxin 10 172 365 modified removed APG00230 Seq ID 271 Alternate start PF00652 Ricin B lectin 35 166 modified PF05431 Toxin 10 176 372 APG00231 Seq ID 272 Alternate start PF14200 RicinB lectin 2 43 146 modified PF05431 Toxin 10 152 349 APG00232 Seq ID 273 Signal peptide PF03945 Endotoxin N 72 272 modified removed PF14200 RicinB lectin 2 522 624 APG00233 Seq ID 274 Signal peptide PF03945 Endotoxin N 47 254 modified removed PF14200 RicinB lectin 2 366 475 APG00235 Seq ID 275 Alternate start PF03945 Endotoxin N 102 320 modified PF14200 RicinB lectin 2 436 539 APG00237 Seq ID 276 Signal peptide PF05431 Toxin 10 166 359 modified removed APG00239 Seq ID 277 Signal peptide PF03945 Endotoxin N 114 337 modified removed PF01473 CW binding 1 394 412 PF01473 CW binding 1 415 434 APG00241 Seq ID 278 Alternate start PF05791 Bacillus HBL 34 215 modified APG00241 Seq ID 279 Signal peptide PF05791 Bacillus HBL 9 190 modified removed APG00242 Seq ID 280 Alternate start PF05431 Toxin 10 193 389 modified APG00243 Seq ID 281 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00248 Seq ID 282 Alternate start PF05431 Toxin 10 191 383 modified APG00249 Seq ID 283 Alternate start PF05431 Toxin 10 210 404 modified APG00249 Seq ID 284 Signal peptide PF05431 Toxin 10 182 376 modified removed APG00251 Seq ID 285 Signal peptide PF03945 Endotoxin N 45 251 modified removed PF14200 RicinB lectin 2 363 472 APG00255 Seq ID 286 Signal peptide PF03945 Endotoxin N 67 299 modified removed PF01473 CW binding 1 346 364 PF14200 RicinB lectin 2 446 549 APG00261 Seq ID 287 Signal peptide PF05431 Toxin 10 169 366 modified removed APG00262 Seq ID 288 Alternate start PF05431 Toxin 10 235 429 modified APG00262 Seq ID 289 Alternate start PF05431 Toxin 10 202 396 modified APG00264 Seq ID 290 Alternate start PF05791 Bacillus HBL 37 217 modified APG00264 Seq ID 291 Signal peptide PF05791 Bacillus HBL 11 191 modified removed APG00266 Seq ID 292 Alternate start PF03945 Endotoxin N 126 319 modified PF01473 CW binding 1 414 433 PF01473 CW binding 1 435 450 APG00267 Seq ID 293 Alternate start PF03945 Endotoxin N 110 309 modified APG00267 Seq ID 294 Signal peptide PF03945 Endotoxin N 72 269 modified removed APG00271 Seq ID 295 Signal peptide PF05791 Bacillus HBL 1 178 modified removed APG00273 Seq ID 296 Alternate start PF12495 Vip3A N 14 186 modified PF02018 CBM 4 9 543 656 APG00274 Seq ID 297 Alternate start PF05791 Bacillus HBL 51 233 modified APG00274 Seq ID 298 Signal peptide PF05791 Bacillus HBL 22 204 modified removed APG00275 Seq ID 299 Alternate start PF03945 Endotoxin N 5 129 modified APG00279 Seq ID 300 Alternate start PF05791 Bacillus HBL 65 181 modified APG00284 Seq ID 302 Alternate start PF14200 RicinB lectin 2 2 98 modified PF05431 Toxin 10 152 349 APG00285 Seq ID 303 Signal peptide PF03945 Endotoxin N 70 300 modified removed PF01473 CW binding 1 369 383 PF01473 CW binding 1 418 432 PF01473 CW binding 1 535 551 APG00286 Seq ID 304 Signal peptide PF05791 Bacillus HBL 12 192 modified removed APG00287 Seq ID 305 Alternate start PF14200 RicinB lectin 2 50 146 modified PF05431 Toxin 10 152 349 APG00288 Seq ID 306 Signal peptide PF03945 Endotoxin N 77 280 modified removed PF01473 CW binding 1 372 391 PF01473 CW binding 1 393 407 APG00290 Seq ID 307 Alternate start PF01338 Bac thur toxin 18 244 modified APG00292 Seq ID 309 Signal peptide PF03945 Endotoxin N 57 266 modified removed PF05588 Botulinum 363 463 HA-17 APG00294 Seq ID 310 Signal peptide PF05791 Bacillus HBL 17 200 modified removed APG00295 Seq ID 311 Alternate start PF05431 Toxin 10 218 414 modified APG00295 Seq ID 312 Signal peptide PF05431 Toxin 10 182 378 modified removed APG00297 Seq ID 313 Alternate start PF05791 Bacillus HBL 36 216 modified APG00297 Seq ID 314 Signal peptide PF05791 Bacillus HBL 10 190 modified removed APG00298 Seq ID 315 Alternate start PF05791 Bacillus HBL 38 218 modified APG00298 Seq ID 316 Signal peptide PF05791 Bacillus HBL 13 193 modified removed APG00301 Seq ID 317 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00302 Seq ID 318 Alternate start PF03945 Endotoxin N 69 278 modified PF14200 RicinB lectin 2 390 485 APG00302 Seq ID 319 Signal peptide PF03945 Endotoxin N 61 270 modified removed PF14200 RicinB lectin 2 382 477 APG00305 Seq ID 320 Signal peptide PF05791 Bacillus HBL 18 204 modified removed APG00307 Seq ID 321 Alternate start PF05791 Bacillus HBL 32 216 modified APG00307 Seq ID 322 Signal peptide PF05791 Bacillus HBL 8 188 modified removed APG00308 Seq ID 323 Signal peptide PF05431 Toxin 10 174 368 modified removed APG00314 Seq ID 324 Alternate start PF05431 Toxin 10 154 348 modified APG00320 Seq ID 325 Alternate start PF05431 Toxin 10 210 404 modified APG00340 Seq ID 326 Alternate start PF14200 RicinB lectin 2 42 143 modified PF05431 Toxin 10 149 347 APG00353 Seq ID 327 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00356 Seq ID 328 Alternate start PF14200 RicinB lectin 2 50 146 modified PF05431 Toxin 10 152 349 APG00368 Seq ID 329 Alternate start PF14200 RicinB lectin 2 2 98 modified PF05431 Toxin 10 152 350 APG00369 Seq ID 330 Signal peptide PF05431 Toxin 10 182 376 modified removed APG00372 Seq ID 331 Signal peptide PF03945 Endotoxin N 80 265 modified removed PF01473 CW binding 1 381 400 PF01473 CW binding 1 402 416 APG00373 Seq ID 332 Alternate start PF00652 Ricin B lectin 37 164 modified PF05431 Toxin 10 175 371 APG00375 Seq ID 333 Signal peptide PF03945 Endotoxin N 79 282 modified removed PF01473 CW binding 1 420 437 APG00376 Seq ID 334 Signal peptide PF05431 Toxin 10 165 357 modified removed APG00377 Seq ID 335 Alternate start PF14200 RicinB lectin 2 43 146 modified PF05431 Toxin 10 152 349 APG00379 Seq ID 336 Alternate start PF14200 RicinB lectin 2 47 148 modified PF05431 Toxin 10 156 353 APG00379 Seq ID 337 Alternate start PF14200 RicinB lectin 2 43 144 modified PF05431 Toxin 10 152 349 APG00385 Seq ID 338 Signal peptide PF05431 Toxin 10 176 369 modified removed APG00391 Seq ID 339 Signal peptide PF03945 Endotoxin N 72 269 modified removed APG00392 Seq ID 340 Signal peptide PF05431 Toxin 10 176 369 modified removed APG00395 Seq ID 341 Signal peptide PF03945 Endotoxin N 70 294 modified removed PF01473 CW binding 1 395 411 PF01473 CW binding 1 423 439 APG00396 Seq ID 342 Signal peptide PF03945 Endotoxin N 76 283 modified removed PF01473 CW binding 1 363 380 PF01473 CW binding 1 382 401 PF01473 CW binding 1 485 500 APG00398 Seq ID 343 Alternate start PF05431 Toxin 10 210 407 modified APG00398 Seq ID 344 Signal peptide PF05431 Toxin 10 182 379 modified removed APG00401 Seq ID 345 Signal peptide PF03945 Endotoxin N 66 298 modified removed PF01473 CW binding 1 345 363 PF14200 RicinB lectin 2 445 548 APG00407 Seq ID 346 Signal peptide PF05431 Toxin 10 181 374 modified removed APG00409 Seq ID 347 Alternate start PF01338 Bac thur toxin 2 198 modified APG00412 Seq ID 348 Alternate start PF05431 Toxin 10 188 381 modified APG00412 Seq ID 349 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00413 Seq ID 350 Alternate start PF00652 Ricin B lectin 35 161 modified PF05431 Toxin 10 171 370 APG00415 Seq ID 351 Alternate start PF05431 Toxin 10 153 346 modified APG00419 Seq ID 352 Signal peptide PF05431 Toxin 10 174 367 modified removed APG00427 Seq ID 353 Signal peptide PF03318 ETX MTX2 52 283 modified removed PF03318 ETX MTX2 52 283 APG00454 Seq ID 354 Alternate start PF05431 Toxin 10 193 389 modified APG00459 Seq ID 355 Signal peptide PF05431 Toxin 10 182 375 modified removed APG00499 Seq ID 356 Alternate start PF00652 Ricin B lectin 24 153 modified PF05431 Toxin 10 164 337 APG00500 Seq ID 357 Alternate start PF03945 Endotoxin N 54 298 modified APG00517 Seq ID 358 Alternate start PF03318 ETX MTX2 51 284 modified APG00521 Seq ID 359 Signal peptide PF05431 Toxin 10 167 360 modified removed APG00559 Seq ID 360 Alternate start PF03318 ETX MTX2 6 224 modified APG00559 Seq ID 361 Alternate start PF03318 ETX MTX2 16 250 modified APG00571 Seq ID 362 Alternate start PF14200 RicinB lectin 2 3 98 modified PF05431 Toxin 10 152 346 APG00590 Seq ID 363 Signal peptide PF03318 ETX MTX2 43 264 modified removed APG00592 Seq ID 364 Alternate start PF05431 Toxin 10 207 400 modified APG00592 Seq ID 365 Signal peptide PF05431 Toxin 10 178 371 modified removed APG00596 Seq ID 366 Alternate start PF00652 Ricin B lectin 37 178 modified PF05431 Toxin 10 179 379 APG00600 Seq ID 367 Signal peptide PF05431 Toxin 10 178 370 modified removed APG00619 Seq ID 368 Signal peptide PF05431 Toxin 10 178 370 modified removed APG00635 Seq ID 369 Alternate start PF03318 ETX MTX2 108 233 modified APG00635 Seq ID 370 Alternate start PF03318 ETX MTX2 85 222 modified APG00635 Seq ID 371 Signal peptide PF03318 ETX MTX2 77 214 modified removed APG00647 Seq ID 372 Alternate start PF01338 Bac thur toxin 17 243 modified APG00652 Seq ID 373 Signal peptide PF05791 Bacillus HBL 10 202 modified removed APG00698 Seq ID 374 Alternate start PF01338 Bac thur toxin 19 248 modified APG00723 Seq ID 375 Alternate start PF03945 Endotoxin N 134 342 modified APG00731 Seq ID 376 Alternate start PF14200 RicinB lectin 2 43 146 modified PF05431 Toxin 10 152 349 APG00748 Seq ID 377 Alternate start PF14200 RicinB lectin 2 42 143 modified PF05431 Toxin 10 149 346 APG00757 Seq ID 378 Alternate start PF00652 Ricin B lectin 39 160 modified PF05431 Toxin 10 170 370 APG00767 Seq ID 379 Signal peptide PF05431 Toxin 10 174 382 modified removed APG00768 Seq ID 380 Signal peptide PF03318 ETX MTX2 70 306 modified removed APG00798 Seq ID 381 Alternate start PF05431 Toxin 10 207 399 modified APG00798 Seq ID 382 Signal peptide PF05431 Toxin 10 178 370 modified removed APG00830 Seq ID 383 Alternate start PF05791 Bacillus HBL 42 233 modified APG00830 Seq ID 384 Signal peptide PF05791 Bacillus HBL 12 203 modified removed APG00844 Seq ID 385 Signal peptide PF05431 Toxin 10 161 354 modified removed APG00851 Seq ID 386 Signal peptide PF03318 ETX MTX2 30 283 modified removed APG00898 Seq ID 387 Alternate start PF03318 ETX MTX2 23 247 modified APG00907 Seq ID 388 Alternate start PF03318 ETX MTX2 26 264 modified APG00913 Seq ID 389 Alternate start PF05431 Toxin 10 210 407 modified APG00925 Seq ID 390 Alternate start PF05431 Toxin 10 209 404 modified APG00568 Seq ID 391 PF14200 RicinB lectin 2 49 150 PF05431 Toxin 10 156 353 APG00568 Seq ID 392 Alternate start PF14200 RicinB lectin 2 45 146 modified PF05431 Toxin 10 152 349 APG00716 Seq ID 393 PF05431 Toxin 10 192 385 APG00716 Seq ID 394 Signal peptide PF05431 Toxin 10 165 358 modified removed APG00736 Seq ID 395 PF01338 Bac thur toxin 100 326 APG00736 Seq ID 396 Alternate start PF01338 Bac thur toxin 19 245 modified APG00930 Seq ID 397 PF03318 ETX MTX2 29 250 APG01245 Seq ID 398 PF03318 ETX MTX2 30 250

Recombinant or synthetic nucleic acids encoding the pesticidal polypeptides disclosed herein are also provided. Of particular interest are nucleic acid sequences that have been designed for expression in a plant of interest. That is, the nucleic acid sequence can be optimized for increased expression in a host plant. A pesticidal protein of the invention can be back-translated to produce a nucleic acid comprising codons optimized for expression in a particular host, for example, a crop plant. In another embodiment, the polynucleotides encoding the polypeptides provided herein may be optimized for increased expression in the transformed plant. That is, the polynucleotides can be synthesized using plant-preferred codons for improved expression. See, for example, Campbell and Gowri (1990) Plant Physiol. 92:1-11 for a discussion of host-preferred codon usage. Methods are available in the art for synthesizing plant-preferred genes. See, for example, U.S. Pat. Nos. 5,380,831, and 5,436,391, and Murray et al. (1989) Nucleic Acids Res. 17:477-498, herein incorporated by reference. Expression of such a coding sequence by the transformed plant (e.g., dicot or monocot) will result in the production of a pesticidal polypeptide and confer increased resistance in the plant to a pest. Recombinant and synthetic nucleic acid molecules encoding the pesticidal proteins of the invention do not include the naturally occurring bacterial sequence encoding the protein.

A “recombinant polynucleotide” or “recombinant nucleic acid” comprises a combination of two or more chemically linked nucleic acid segments which are not found directly joined in nature. By “directly joined” is intended the two nucleic acid segments are immediately adjacent and joined to one another by a chemical linkage. In specific embodiments, the recombinant polynucleotide comprises a polynucleotide of interest or a variant or fragment thereof such that an additional chemically linked nucleic acid segment is located either 5′, 3′ or internal to the polynucleotide of interest. Alternatively, the chemically-linked nucleic acid segment of the recombinant polynucleotide can be formed by deletion of a sequence. The additional chemically linked nucleic acid segment or the sequence deleted to join the linked nucleic acid segments can be of any length, including for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20 or greater nucleotides.

Various methods for making such recombinant polynucleotides include chemical synthesis or by the manipulation of isolated segments of polynucleotides by genetic engineering techniques. In specific embodiments, the recombinant polynucleotide can comprise a recombinant DNA sequence or a recombinant RNA sequence. A “fragment of a recombinant polynucleotide or nucleic acid” comprises at least one of a combination of two or more chemically linked amino acid segments which are not found directly joined in nature.

Fragments of a polynucleotide (RNA or DNA) may encode protein fragments that retain activity. In specific embodiments, a fragment of a recombinant polynucleotide or a recombinant polynucleotide construct comprises at least one junction of the two or more chemically linked or operably linked nucleic acid segments which are not found directly joined in nature. A fragment of a polynucleotide that encodes a biologically active portion of a polypeptide that retains pesticidal activity will encode at least 25, 30, 40, 50, 60, 70, 75, 80, 90, 100, 110, 120, 125, 130, 140, 150, 160, 170, 175, 180, contiguous amino acids, or up to the total number of amino acids present in a full-length polypeptide as set forth in SEQ ID NO:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398. In specific embodiments, such polypeptide fragments are active fragments, and in still other embodiments, the polypeptide fragment comprises a recombinant polypeptide fragment. As used herein, a fragment of a recombinant polypeptide comprises at least one of a combination of two or more chemically linked amino acid segments which are not found directly joined in nature.

By “variants” is intended to mean substantially similar sequences. For polynucleotides, a variant comprises a deletion and/or addition of one or more nucleotides at one or more internal sites within the native polynucleotide and/or a substitution of one or more nucleotides at one or more sites in the native polynucleotide. As used herein, a “native” polynucleotide or polypeptide comprises a naturally occurring nucleotide sequence or amino acid sequence, respectively.

Variants of a particular polynucleotide of the invention (i.e., the reference polynucleotide) can also be evaluated by comparison of the percent sequence identity between the polypeptide encoded by a variant polynucleotide and the polypeptide encoded by the reference polynucleotide. Thus, for example, an isolated polynucleotide that encodes a polypeptide with a given percent sequence identity to the polypeptide of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398 are disclosed. Percent sequence identity between any two polypeptides can be calculated using sequence alignment programs and parameters described elsewhere herein. Where any given pair of polynucleotides of the invention is evaluated by comparison of the percent sequence identity shared by the two polypeptides they encode, the percent sequence identity between the two encoded polypeptides is at least about 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398. In other embodiments, the variant of the polynucleotide provided herein differs from the native sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more nucleotides.

Variant polynucleotide and proteins also encompass sequences and proteins derived from a mutagenic and recombinogenic procedure such as DNA shuffling. With such a procedure, one or more different pesticidal protein disclosed herein (SEQ ID NO: 1-209) is manipulated to create a new pesticidal protein possessing the desired properties. In this manner, libraries of recombinant polynucleotides are generated from a population of related sequence polynucleotides comprising sequence regions that have substantial sequence identity and can be homologously recombined in vitro or in vivo. For example, using this approach, sequence motifs encoding a domain of interest may be shuffled between the pesticidal sequences provided herein and other known pesticidal genes to obtain a new gene coding for a protein with an improved property of interest, such as an increased K_(m) in the case of an enzyme. Strategies for such DNA shuffling are known in the art. See, for example, Stemmer (1994) Proc. Natl. Acad. Sci. USA 91:10747-10751; Stemmer (1994) Nature 370:389-391; Crameri et al. (1997) Nature Biotech. 15:436-438; Moore et al. (1997) J. Mol. Biol. 272:336-347; Zhang et al. (1997) Proc. Natl. Acad. Sci. USA 94:4504-4509; Crameri et al. (1998) Nature 391:288-291; and U.S. Pat. Nos. 5,605,793 and 5,837,458. A “shuffled” nucleic acid is a nucleic acid produced by a shuffling procedure such as any shuffling procedure set forth herein. Shuffled nucleic acids are produced by recombining (physically or virtually) two or more nucleic acids (or character strings), for example in an artificial, and optionally recursive, fashion. Generally, one or more screening steps are used in shuffling processes to identify nucleic acids of interest; this screening step can be performed before or after any recombination step. In some (but not all) shuffling embodiments, it is desirable to perform multiple rounds of recombination prior to selection to increase the diversity of the pool to be screened. The overall process of recombination and selection are optionally repeated recursively. Depending on context, shuffling can refer to an overall process of recombination and selection, or, alternately, can simply refer to the recombinational portions of the overall process.

In one embodiment, a method of obtaining a polynucleotide that encodes an improved polypeptide comprising pesticidal activity is provided, wherein the improved polypeptide has at least one improved property over any one of SEQ ID NOS: 1-229. Such methods can comprise (a) recombining a plurality of parental polynucleotides to produce a library of recombinant polynucleotides encoding recombinant pesticidal polypeptides; (b) screening the library to identify a recombinant polynucleotide that encodes an improved recombinant pesticidal polypeptide that has an enhanced property improved over the parental polynucleotide; (c) recovering the recombinant polynucleotide that encodes the improved recombinant pesticidal polypeptide identified in (b); and, (d) repeating steps (a), (b) and (c) using the recombinant polynucleotide recovered in step (c) as one of the plurality of parental polynucleotides in repeated step (a).

iii. Sequence Comparisons

As used herein, the term “identity” or “percent identity” when used with respect to a particular pair of aligned amino acid sequences, refers to the percent amino acid sequence identity that is obtained by counting the number of identical matches in the alignment and dividing such number of identical matches by the length of the aligned sequences. As used herein, the term “similarity” or “percent similarity” when used with respect to a particular pair of aligned amino acid sequences, refers to the sum of the scores that are obtained from a scoring matrix for each amino acid pair in the alignment divided by the length of the aligned sequences.

Unless otherwise stated, identity and similarity will be calculated by the Needleman-Wunsch global alignment and scoring algorithms (Needleman and Wunsch (1970) J. Mol. Biol. 48(3):443-453) as implemented by the “needle” program, distributed as part of the EMBOSS software package (Rice, P. Longden, I. and Bleasby, A., EMBOSS: The European Molecular Biology Open Software Suite, 2000, Trends in Genetics 16, (6) pp 276-277, versions 6.3.1 available from EMBnet at embnet.org/resource/emboss and emboss.sourceforge.net, among other sources) using default gap penalties and scoring matrices (EBLOSUM62 for protein and EDNAFULL for DNA). Equivalent programs may also be used. By “equivalent program” is intended any sequence comparison program that, for any two sequences in question, generates an alignment having identical nucleotide residue matches and an identical percent sequence identity when compared to the corresponding alignment generated by needle from EMBOSS version 6.3.1.

Additional mathematical algorithms are known in the art and can be utilized for the comparison of two sequences. See, for example, the algorithm of Karlin and Altschul (1990) Proc. Natl. Acad. Sci. USA 87:2264, modified as in Karlin and Altschul (1993) Proc. Natl. Acad. Sci. USA 90:5873-5877. Such an algorithm is incorporated into the BLAST programs of Altschul et al. (1990) J. Mol. Biol. 215:403. BLAST nucleotide searches can be performed with the BLASTN program (nucleotide query searched against nucleotide sequences) to obtain nucleotide sequences homologous to pesticidal-like nucleic acid molecules of the invention, or with the BLASTX program (translated nucleotide query searched against protein sequences) to obtain protein sequences homologous to pesticidal nucleic acid molecules of the invention. BLAST protein searches can be performed with the BLASTP program (protein query searched against protein sequences) to obtain amino acid sequences homologous to pesticidal protein molecules of the invention, or with the TBLASTN program (protein query searched against translated nucleotide sequences) to obtain nucleotide sequences homologous to pesticidal protein molecules of the invention. To obtain gapped alignments for comparison purposes, Gapped BLAST (in BLAST 2.0) can be utilized as described in Altschul et al. (1997) Nucleic Acids Res. 25:3389. Alternatively, PSI-Blast can be used to perform an iterated search that detects distant relationships between molecules. See Altschul et al. (1997) supra. When utilizing BLAST, Gapped BLAST, and PSI-Blast programs, the default parameters of the respective programs (e.g., BLASTX and BLASTN) can be used. Alignment may also be performed manually by inspection.

Two sequences are “optimally aligned” when they are aligned for similarity scoring using a defined amino acid substitution matrix (e.g., BLOSUM62), gap existence penalty and gap extension penalty so as to arrive at the highest score possible for that pair of sequences. Amino acid substitution matrices and their use in quantifying the similarity between two sequences are well-known in the art and described, e.g., in Dayhoff et al. (1978) “A model of evolutionary change in proteins.” In “Atlas of Protein Sequence and Structure,” Vol. 5, Suppl. 3 (ed. M. O. Dayhoff), pp. 345-352. Natl. Biomed. Res. Found., Washington, D.C. and Henikoff et al. (1992) Proc. Natl. Acad. Sci. USA 89:10915-10919. The BLOSUM62 matrix is often used as a default scoring substitution matrix in sequence alignment protocols. The gap existence penalty is imposed for the introduction of a single amino acid gap in one of the aligned sequences, and the gap extension penalty is imposed for each additional empty amino acid position inserted into an already opened gap. The alignment is defined by the amino acids positions of each sequence at which the alignment begins and ends, and optionally by the insertion of a gap or multiple gaps in one or both sequences, so as to arrive at the highest possible score. While optimal alignment and scoring can be accomplished manually, the process is facilitated by the use of a computer-implemented alignment algorithm, e.g., gapped BLAST 2.0, described in Altschul et al. (1997) Nucleic Acids Res. 25:3389-3402, and made available to the public at the National Center for Biotechnology Information Website. Optimal alignments, including multiple alignments, can be prepared using, e.g., PSI-BLAST, and described by Altschul et al. (1997) Nucleic Acids Res. 25:3389-3402.

With respect to an amino acid sequence that is optimally aligned with a reference sequence, an amino acid residue “corresponds to” the position in the reference sequence with which the residue is paired in the alignment. The “position” is denoted by a number that sequentially identifies each amino acid in the reference sequence based on its position relative to the N-terminus. For example, in SEQ ID NO: 177 position 1 is M, position 2 is N, position 3 is E, etc. When a test sequence is optimally aligned with SEQ ID NO: 177, a residue in the test sequence that aligns with the E at position 3 is said to “correspond to position 3” of SEQ ID NO: 177. Owing to deletions, insertion, truncations, fusions, etc., that must be taken into account when determining an optimal alignment, in general the amino acid residue number in a test sequence as determined by simply counting from the N-terminal will not necessarily be the same as the number of its corresponding position in the reference sequence. For example, in a case where there is a deletion in an aligned test sequence, there will be no amino acid that corresponds to a position in the reference sequence at the site of deletion. Where there is an insertion in an aligned reference sequence, that insertion will not correspond to any amino acid position in the reference sequence. In the case of truncations or fusions there can be stretches of amino acids in either the reference or aligned sequence that do not correspond to any amino acid in the corresponding sequence.

iv. Antibodies

Antibodies to the polypeptides of the present invention, or to variants or fragments thereof, are also encompassed. Methods for producing antibodies are well known in the art (see, for example, Harlow and Lane (1988) Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.; and U.S. Pat. No. 4,196,265). These antibodies can be used in kits for the detection and isolation of toxin polypeptides. Thus, this disclosure provides kits comprising antibodies that specifically bind to the polypeptides described herein, including, for example, polypeptides having the sequence of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398.

II. Pests

The compositions and methods provided herein are useful against a variety of pests. “Pests” includes but is not limited to, insects, fungi, bacteria, nematodes, acarids, protozoan pathogens, animal-parasitic liver flukes, and the like. Pests of particular interest are insect pests, particularly insect pests that cause significant damage to agricultural plants. Insect pests include insects selected from the orders Coleoptera, Diptera, Hymenoptera, Lepidoptera, Mallophaga, Homoptera, Hemiptera, Orthroptera, Thysanoptera, Dermaptera, Isoptera, Anoplura, Siphonaptera, Trichoptera, or nematodes. In non-limiting embodiments, the insect pest comprises Western corn rootworm, Diabrotica virgifera virgifera; Fall armyworm, Spodoptera frupperda; Colorado potato beetle, Leptinotarsa decemlineata; Corn earworm, Helicoverpa zea (in North America same species attacks cotton and called cotton bollworm); European corn borer, Ostrinia nubilalis; Black cutworm, Agrotis Ipsilon; Diamondback moth, Plutella xylostella; Velvetbean caterpillar, Anticarsia gemmatalis; Southwestern corn borer, Diatraea grandiosealla; Cotton bollworm, Helicoverpa armigera (found other than USA in rest of the world); Southern green stinkbug, Nezara viridula; Green stinkbug, Chinavia halaris; Brown marmorated stinkbug, Halyomorpha halys; and Brown stinbug, Euschistus servus Euschistus heros (Neotropical brown stink bug OR soy stink bug); Piezodorus guildinii (red-banded stink bug); Dichelops melacanthus (no common name) and/or Dichelops furcatus (no common name); an aphid, such as a soybean aphid. In other embodiments, the pest comprises a nematode including, but not limited to, Meloidogyne hapla (Northern root-knot nematode); Meloidogyne enterolobii, Meloidogyne arenaria (peanut root-knot nematode); and Meloidogyne javanica.

The term “insect pests” as used herein refers to insects and other similar pests such as, for example, those of the order Acari including, but not limited to, mites and ticks. Insect pests of the present invention include, but are not limited to, insects of the order Lepidoptera, e.g. Achoroia grisella, Acleris gloverana, Acleris variana, Adoxophyes orana, Agrotis ipsilon, Alabama argillacea, Alsophila pometaria, Amyelois transitella, Anagasta kuehniella, Anarsia lineatella, Anisota senatoria, Antheraea pernyi, Anticarsia gemmatalis, Archips sp., Argyrotaenia sp., Athetis mindara, Bombyx mori, Bucculatrix thurberiella, Cadra cautella, Choristoneura sp., Cochylls hospes, Colias eurytheme, Corcyra cephalonica, Cydia latiferreanus, Cydia pomonella, Datana integerrima, Dendrolimus sibericus, Desmiafeneralis, Diaphania hyalinata, Diaphania nitidalis, Diatraea grandiosella, Diatraea saccharalis, Ennomos subsignaria, Eoreuma loftini, Esphestia elutella, Erannis tilaria, Estigmene acrea, Eulia salubricola, Eupocoellia ambiguella, Eupoecilia ambiguella, Euproctis chrysorrhoea, Euxoa messoria, Galleria mellonella, Grapholita molesta, Harrisina americana, Helicoverpa subflexa, Helicoverpa zea, Heliothis virescens, Hemileuca oliviae, Homoeosoma electellum, Hyphantia cunea, Keiferia lycopersicella, Lambdina fiscellaria fiscellaria, Lambdina fiscellaria lugubrosa, Leucoma salicis, Lobesia botrana, Loxostege sticticalis, Lymantria dispar, Macalla thyrisalis, Malacosoma sp., Mamestra brassicae, Mamestra configurata, Manduca quinquemaculata, Manduca sexta, Maruca testulalis, Melanchra picta, Operophtera brumata, Orgyia sp., Ostrinia nubilalis, Paleacrita vernata, Papilio cresphontes, Pectinophora gossypiella, Phryganidia californica, Phyllonorycter blancardella, Pieris napi, Pieris rapae, Plathypena scabra, Platynota flouendana, Platynota stultana, Platyptilia carduidactyla, Plodia interpunctella, Plutella xylostella, Pontia protodice, Pseudaletia unipuncta, Pseudoplasia includens, Sabulodes aegrotata, Schizura concinna, Sitotroga cerealella, Spilonta ocellana, Spodoptera sp., Thaurnstopoea pityocampa, Tinsola bisselliella, Trichoplusia hi, Udea rubigalis, Xylomyges curiails, and Yponomeuta padella.

Insect pests also include insects selected from the orders Diptera, Hymenoptera, Lepidoptera, Mallophaga, Homoptera, Hemiptera, Orthroptera, Thysanoptera, Dermaptera, Isoptera, Anoplura, Siphonaptera, Trichoptera, Coleoptera.

Insect pests of the invention for the major crops include, but are not limited to: Maize: Ostrinia nubilalis, European corn borer; Agrotis ipsilon, black cutworm; Helicoverpa zeae, corn earworm; Spodoptera frugiperda, fall armyworm; Diatraea grandiosella, southwestern corn borer; Elasmopalpus lignosellus, lesser cornstalk borer; Diatraea saccharalis, surgarcane borer; western corn rootworm, e.g., Diabrotica virgifera virgifera; northern corn rootworm, e.g., Diabrotica longicornis barberi; southern corn rootworm, e.g., Diabrotica undecimpunctata howardi; Melanotus spp., wireworms; Cyclocephala borealis, northern masked chafer (white grub); Cyclocephala immaculata, southern masked chafer (white grub); Popillia japonica, Japanese beetle; Chaetocnema pulicaria, corn flea beetle; Sphenophorus maidis, maize billbug; Rhopalosiphum maidis, corn leaf aphid; Anuraphis maidiradicis, corn root aphid; Blissus leucopterus leucopterus, chinch bug; Melanoplus femurrubrum, redlegged grasshopper; Melanoplus sanguinipes, migratory grasshopper; Hylemya platura, seedcorn maggot; Agromyza parvicornis, corn blotch leafminer; Anaphothrips obscrurus, grass thrips; Solenopsis milesta, thief ant; Tetranychus urticae, two spotted spider mite; Sorghum: Chilo partellus, sorghum borer; Spodoptera frugiperda, fall armyworm; Helicoverpa zea, corn earworm; Elasmopalpus lignosellus, lesser cornstalk borer; Feltia subterranea, granulate cutworm; Phyllophaga crinita, white grub; Eleodes, Conoderus, and Aeolus spp., wireworms; Oulema melanopus, cereal leaf beetle; Chaetocnema pulicaria, corn flea beetle; Sphenophorus maidis, maize billbug; Rhopalosiphum maidis; corn leaf aphid; Sipha flava, yellow sugarcane aphid; chinch bug, e.g., Blissus leucopterus leucopterus; Contarinia sorghicola, sorghum midge; Tetranychus cinnabarinus, carmine spider mite; Tetranychus urticae, two-spotted spider mite; Wheat: Pseudaletia unipunctata, army worm; Spodoptera frugiperda, fall armyworm; Elasmopalpus lignosellus, lesser cornstalk borer; Agrotis orthogonia, pale western cutworm; Elasmopalpus lignosellus, lesser cornstalk borer; Oulema melanopus, cereal leaf beetle; Hypera punctata, clover leaf weevil; southern corn rootworm, e.g., Diabrotica undecimpunctata howardi; Russian wheat aphid; Schizaphis graminum, greenbug; Macrosiphum avenae, English grain aphid; Melanoplus femurrubrum, redlegged grasshopper; Melanoplus differentialis, differential grasshopper; Melanoplus sanguinipes, migratory grasshopper; Mayetiola destructor, Hessian fly; Sitodiplosis mosellana, wheat midge; Meromyza americana, wheat stem maggot; Hylemya coarctata, wheat bulb fly; Frankliniella fusca, tobacco thrips; Cephus cinctus, wheat stem sawfly; Aceria tulipae, wheat curl mite; Sunflower: Cylindrocupturus adspersus, sunflower stem weevil; Smicronyx fulus, red sunflower seed weevil; Smicronyx sordidus, gray sunflower seed weevil; Suleima helianthana, sunflower bud moth; Homoeosoma electellum, sunflower moth; Zygogramma exclamationis, sunflower beetle; Bothyrus gibbosus, carrot beetle; Neolasioptera murtfeldtiana, sunflower seed midge; Cotton: Heliothis virescens, tobacco budworm; Helicoverpa zea, cotton bollworm; Spodoptera exigua, beet armyworm; Pectinophora gossypiella, pink bollworm; boll weevil, e.g., Anthonomus grandis; Aphis gossypii, cotton aphid; Pseudatomoscelis seriatus, cotton fleahopper; Trialeurodes abutilonea, bandedwinged whitefly; Lygus lineolaris, tarnished plant bug; Melanoplus femurrubrum, redlegged grasshopper; Melanoplus differentialis, differential grasshopper; Thrips tabaci, onion thrips; Franklinkiella fusca, tobacco thrips; Tetranychus cinnabarinus, carmine spider mite; Tetranychus urticae, two-spotted spider mite; Rice: Diatraea saccharalis, sugarcane borer; Spodoptera frupperda, fall armyworm; Helicoverpa zea, corn earworm; Colaspis brunnea, grape colaspis; Lissorhoptrus oryzophilus, rice water weevil; Sitophilus oryzae, rice weevil; Nephotettix nigropictus, rice leafhoper; chinch bug, e.g., Blissus leucopterus leucopterus; Acrosternum hilare, green stink bug; Soybean: Pseudoplusia includens, soybean looper; Anticarsia gemmatalis, velvetbean caterpillar; Plathypena scabra, green cloverworm; Ostrinia nubilalis, European corn borer; Agrotis ipsilon, black cutworm; Spodoptera exigua, beet armyworm; Heliothis virescens, tobacco budworm; Helicoverpa zea, cotton bollworm; Epilachna varivestis, Mexican bean beetle; Myzus persicae, green peach aphid; Empoasca fabae, potato leafhopper; Acrosternum hilare, green stink bug; Melanoplus femurrubrum, redlegged grasshopper; Melanoplus differentialis, differential grasshopper; Hylemya platura, seedcorn maggot; Sericothrips variabilis, soybean thrips; Thrips tabaci, onion thrips; Tetranychus turkestani, strawberry spider mite; Tetranychus urticae, two-spotted spider mite; Barley: Ostrinia nubilalis, European corn borer; Agrotis ipsilon, black cutworm; Schizaphis graminum, greenbug; chinch bug, e.g., Blissus leucopterus leucopterus; Acrosternum hilare, green stink bug; Euschistus servus, brown stink bug; Jylemya platura, seedcorn maggot; Mayetiola destructor, Hessian fly; Petrobia latens, brown wheat mite; Oil Seed Rape: Vrevicoryne brassicae, cabbage aphid; Phyllotreta cruciferae, crucifer flea beetle; Phyllotreta striolata, striped flea beetle; Phyllotreta nemorum, striped turnip flea beetle; Meligethes aeneus, rapeseed beetle; and the pollen beetles Meligethes rufimanus, Meligethes nigrescens, Meligethes canadianus, and Meligethes viridescens; Potato: Leptinotarsa decemlineata, Colorado potato beetle.

The methods and compositions provided herein may be effective against Hemiptera such as Lygus hesperus, Lygus lineolaris, Lygus pratensis, Lygus ruguhpennis Popp, Lygus pabulinus, Calocoris norvegicus, Orthops compestris, Plesiocoris rugicollis, Cyrtopeltis modestus, Cyrtopeltis notatus, Spanagonicus albofasciatus, Diaphnocoris chlorinonis, Labopidicola allii, Pseudatomoscelis seriatus, Adelphocoris rapidus, Poecilocapsus lineatus, Blissus leucopterus, Nysius ericae, Nysius raphanus, Euschistus servus, Nezara viridula, Eurygaster, Coreidae, Pyrrhocoridae, Tinidae, Blostomatidae, Reduviidae, and Cimicidae. Pests of interest also include Araecerus fasciculatus, coffee bean weevil; Acanthoscelides obtectus, bean weevil; Bruchus rufmanus, broadbean weevil; Bruchus pisorum, pea weevil; Zabrotes subfasciatus, Mexican bean weevil; Diabrotica balteata, banded cucumber beetle; Cerotoma trifurcata, bean leaf beetle; Diabrotica virgifera, Mexican corn rootworm; Epitrix cucumeris, potato flea beetle; Chaetocnema confinis, sweet potato flea beetle; Hypera postica, alfalfa weevil; Anthonomus quadrigibbus, apple curculio; Sternechus paludatus, bean stalk weevil; Hypera brunnipennis, Egyptian alfalfa weevil; Sitophilus granaries, granary weevil; Craponius inaequalis, grape curculio; Sitophilus zeamais, maize weevil; Conotrachelus nenuphar, plum curculio; Euscepes postfaciatus, West Indian sweet potato weevil; Maladera castanea, Asiatic garden beetle; Rhizotrogus majalis, European chafer; Macrodactylus subspinosus, rose chafer; Tribolium confusum, confused flour beetle; Tenebrio obscurus, dark mealworm; Tribolium castaneum, red flour beetle; Tenebrio molitor, yellow mealworm.

Nematodes include parasitic nematodes such as root-knot, cyst, and lesion nematodes, including Heterodera spp., Meloidogyne spp., and Globodera spp.; particularly members of the cyst nematodes, including, but not limited to, Heterodera glycines (soybean cyst nematode); Heterodera schachtii (beet cyst nematode); Heterodera avenae (cereal cyst nematode); and Globodera rostochiensis and Globodera pailida (potato cyst nematodes). Lesion nematodes include Pratylenchus spp.

Insect pests may be tested for pesticidal activity of compositions of the invention in early developmental stages, e.g., as larvae or other immature forms. The insects may be reared in total darkness at from about 20° C. to about 30° C. and from about 30% to about 70% relative humidity. Bioassays may be performed as described in Czapla and Lang (1990) J. Econ. Entomol. 83 (6): 2480-2485. See, also the experimental section herein.

III. Expression Cassettes

Polynucleotides encoding the pesticidal proteins provided herein can be provided in expression cassettes for expression in an organism of interest. The cassette will include 5′ and 3′ regulatory sequences operably linked to a polynucleotide encoding a pesticidal polypeptide provided herein that allows for expression of the polynucleotide. The cassette may additionally contain at least one additional gene or genetic element to be cotransformed into the organism. Where additional genes or elements are included, the components are operably linked. Alternatively, the additional gene(s) or element(s) can be provided on multiple expression cassettes. Such an expression cassette is provided with a plurality of restriction sites and/or recombination sites for insertion of the polynucleotides to be under the transcriptional regulation of the regulatory regions. The expression cassette may additionally contain a selectable marker gene.

The expression cassette will include in the 5′-3′ direction of transcription, a transcriptional and translational initiation region (i.e., a promoter), a pesticidal polynucleotide of the invention, and a transcriptional and translational termination region (i.e., termination region) functional in the organism of interest, i.e., a plant or bacteria. The promoters of the invention are capable of directing or driving expression of a coding sequence in a host cell. The regulatory regions (i.e., promoters, transcriptional regulatory regions, and translational termination regions) may be endogenous or heterologous to the host cell or to each other. As used herein, “heterologous” in reference to a sequence is a sequence that originates from a foreign species, or, if from the same species, is substantially modified from its native form in composition and/or genomic locus by deliberate human intervention. As used herein, a chimeric gene comprises a coding sequence operably linked to a transcription initiation region that is heterologous to the coding sequence.

Convenient termination regions are available from the Ti-plasmid of A. tumefaciens, such as the octopine synthase and nopaline synthase termination regions. See also Guerineau et al. (1991) Mol. Gen. Genet. 262:141-144; Proudfoot (1991) Cell 64:671-674; Sanfacon et al. (1991) Genes Dev. 5:141-149; Mogen et al. (1990) Plant Cell 2:1261-1272; Munroe et al. (1990) Gene 91:151-158; Ballas et al. (1989) Nucleic Acids Res. 17:7891-7903; and Joshi et al. (1987) Nucleic Acids Res. 15:9627-9639.

Additional regulatory signals include, but are not limited to, transcriptional initiation start sites, operators, activators, enhancers, other regulatory elements, ribosomal binding sites, an initiation codon, termination signals, and the like. See, for example, U.S. Pat. Nos. 5,039,523 and 4,853,331; EPO 0480762A2; Sambrook et al. (1992) Molecular Cloning: A Laboratory Manual, ed. Maniatis et al. (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.), hereinafter “Sambrook 11”; Davis et al., eds. (1980) Advanced Bacterial Genetics (Cold Spring Harbor Laboratory Press), Cold Spring Harbor, N.Y., and the references cited therein.

In preparing the expression cassette, the various DNA fragments may be manipulated, so as to provide for the DNA sequences in the proper orientation and, as appropriate, in the proper reading frame. Toward this end, adapters or linkers may be employed to join the DNA fragments or other manipulations may be involved to provide for convenient restriction sites, removal of superfluous DNA, removal of restriction sites, or the like. For this purpose, in vitro mutagenesis, primer repair, restriction, annealing, resubstitutions, e.g., transitions and transversions, may be involved.

A number of promoters can be used in the practice of the invention. The promoters can be selected based on the desired outcome. The nucleic acids can be combined with constitutive, inducible, tissue-preferred, or other promoters for expression in the organism of interest. See, for example, promoters set forth in WO 99/43838 and in U.S. Pat. Nos. 8,575,425; 7,790,846; 8,147,856; 8,586832; 7,772,369; 7,534,939; 6,072,050; 5,659,026; 5,608,149; 5,608,144; 5,604,121; 5,569,597; 5,466,785; 5,399,680; 5,268,463; 5,608,142; and 6,177,611; herein incorporated by reference.

For expression in plants, constitutive promoters also include CaMV 35S promoter (Odell et al. (1985) Nature 313:810-812); rice actin (McElroy et al. (1990) Plant Cell 2:163-171); ubiquitin (Christensen et al. (1989) Plant Mol. Biol. 12:619-632 and Christensen et al. (1992) Plant Mol. Biol. 18:675-689); pEMU (Last et al. (1991) Theor. Appl. Genet. 81:581-588); MAS (Velten et al. (1984) EMBO J. 3:2723-2730). Inducible promoters include those that drive expression of pathogenesis-related proteins (PR proteins), which are induced following infection by a pathogen. See, for example, Redolfi et al. (1983) Neth. J. Plant Pathol. 89:245-254; Uknes et al. (1992) Plant Cell 4:645-656; and Van Loon (1985) Plant Mol. Virol. 4:111-116; and WO 99/43819, herein incorporated by reference. Promoters that are expressed locally at or near the site of pathogen infection may also be used (Marineau et al. (1987) Plant Mol. Biol. 9:335-342; Matton et al. (1989) Molecular Plant-Microbe Interactions 2:325-331; Somsisch et al. (1986) Proc. Natl. Acad. Sci. USA 83:2427-2430; Somsisch et al. (1988) Mol. Gen. Genet. 2:93-98; and Yang (1996) Proc. Natl. Acad. Sci. USA 93:14972-14977; Chen et al. (1996) Plant J. 10:955-966; Zhang et al. (1994) Proc. Natl. Acad. Sci. USA 91:2507-2511; Warner et al. (1993) Plant J. 3:191-201; Siebertz et al. (1989) Plant Cell 1:961-968; Cordero et al. (1992) Physiol. Mol. Plant Path. 41:189-200; U.S. Pat. No. 5,750,386 (nematode-inducible); and the references cited therein).

Wound-inducible promoters may be used in the constructions of the invention. Such wound-inducible promoters include pin II promoter (Ryan (1990) Ann. Rev. Phytopath. 28:425-449; Duan et al. (1996) Nature Biotechnology 14:494-498); wun1 and wun2 (U.S. Pat. No. 5,428,148); win1 and win2 (Stanford et al. (1989) Mol. Gen. Genet. 215:200-208); systemin (McGurl et al. (1992) Science 225:1570-1573); WIP1 (Rohmeier et al. (1993) Plant Mol. Biol. 22:783-792; Eckelkamp et al. (1993) FEBS Letters 323:73-76); MPI gene (Corderok et al. (1994) Plant J. 6(2):141-150); and the like, herein incorporated by reference.

Tissue-preferred promoters for use in the invention include those set forth in Yamamoto et al. (1997) Plant J. 12(2):255-265; Kawamata et al. (1997) Plant Cell Physiol. 38(7):792-803; Hansen et al. (1997) Mol. Gen Genet. 254(3):337-343; Russell et al. (1997) Transgenic Res. 6(2):157-168; Rinehart et al. (1996) Plant Physiol. 112(3):1331-1341; Van Camp et al. (1996) Plant Physiol. 112(2):525-535; Canevascini et al. (1996) Plant Physiol. 112(2):513-524; Yamamoto et al. (1994) Plant Cell Physiol. 35(5):773-778; Lam (1994) Results Probl. Cell Differ. 20:181-196; Orozco et al. (1993) Plant Mol Biol. 23(6):1129-1138; Matsuoka et al. (1993) Proc Natl. Acad. Sci. USA 90(20):9586-9590; and Guevara-Garcia et al. (1993) Plant J. 4(3):495-505.

Leaf-preferred promoters include those set forth in Yamamoto et al. (1997) Plant J. 12(2):255-265; Kwon et al. (1994) Plant Physiol. 105:357-67; Yamamoto et al. (1994) Plant Cell Physiol. 35(5):773-778; Gotor et al. (1993) Plant J. 3:509-18; Orozco et al. (1993) Plant Mol. Biol. 23(6):1129-1138; and Matsuoka et al. (1993) Proc. Natl. Acad. Sci. USA 90(20):9586-9590.

Root-preferred promoters are known and include those in Hire et al. (1992) Plant Mol. Biol. 20(2):207-218 (soybean root-specific glutamine synthetase gene); Keller and Baumgartner (1991) Plant Cell 3(10):1051-1061 (root-specific control element); Sanger et al. (1990) Plant Mol. Biol. 14(3):433-443 (mannopine synthase (MAS) gene of Agrobacterium tumefaciens); and Miao et al. (1991) Plant Cell 3(1):11-22 (cytosolic glutamine synthetase (GS)); Bogusz et al. (1990) Plant Cell 2(7):633-641; Leach and Aoyagi (1991) Plant Science (Limerick) 79(1):69-76 (rolC and rolD); Teeri et al. (1989) EMBO J. 8(2):343-350; Kuster et al. (1995) Plant Mol. Biol. 29(4):759-772 (the VfENOD-GRP3 gene promoter); and, Capana et al. (1994) Plant Mol. Biol. 25(4):681-691 (rolB promoter). See also U.S. Pat. Nos. 5,837,876; 5,750,386; 5,633,363; 5,459,252; 5,401,836; 5,110,732; and 5,023,179.

“Seed-preferred” promoters include both “seed-specific” promoters (those promoters active during seed development such as promoters of seed storage proteins) as well as “seed-germinating” promoters (those promoters active during seed germination). See Thompson et al. (1989) BioEssays 10:108. Seed-preferred promoters include, but are not limited to, Cim1 (cytokinin-induced message); cZ19B1 (maize 19 kDa zein); mi1ps (myo-inositol-1-phosphate synthase) (see WO 00/11177 and U.S. Pat. No. 6,225,529). Gamma-zein is an endosperm-specific promoter. Globulin 1 (Glb-1) is a representative embryo-specific promoter. For dicots, seed-specific promoters include, but are not limited to, bean β-phaseolin, napin, β-conglycinin, soybean lectin, cruciferin, and the like. For monocots, seed-specific promoters include, but are not limited to, maize 15 kDa zein, 22 kDa zein, 27 kDa zein, gamma-zein, waxy, shrunken 1, shrunken 2, Globulin 1, etc. See also WO 00/12733, where seed-preferred promoters from end1 and end2 genes are disclosed.

For expression in a bacterial host, promoters that function in bacteria are well-known in the art. Such promoters include any of the known crystal protein gene promoters, including the promoters of any of the pesticidal proteins of the invention, and promoters specific for B. thuringiensis sigma factors. Alternatively, mutagenized or recombinant crystal protein-encoding gene promoters may be recombinantly engineered and used to promote expression of the novel gene segments disclosed herein.

The expression cassette can also comprise a selectable marker gene for the selection of transformed cells. Selectable marker genes are utilized for the selection of transformed cells or tissues. Marker genes include genes encoding antibiotic resistance, such as those encoding neomycin phosphotransferase II (NEO) and hygromycin phosphotransferase (HPT), as well as genes conferring resistance to herbicidal compounds, such as glufosinate ammonium, bromoxynil, imidazolinones, and 2,4-dichlorophenoxyacetate (2,4-D). Additional selectable markers are known and any can be used in the practice of the invention. See, for example, PCT/US2015/066648, filed on Dec. 18, 2015, herein incorporated by reference in its entirety, which discloses glufosinate resistance sequences that can be employed as selectable markers.

IV. Methods, Host Cells and Plant Cells

As indicated, DNA constructs comprising nucleotide sequences encoding the pesticidal proteins or active variants or fragment thereof can be used to transform plants of interest or other organisms of interest. Methods for transformation involve introducing a nucleotide construct into a plant. By “introducing” is intended to introduce the nucleotide construct to the plant or other host cell in such a manner that the construct gains access to the interior of a cell of the plant or host cell. The methods of the invention do not require a particular method for introducing a nucleotide construct to a plant or host cell, only that the nucleotide construct gains access to the interior of at least one cell of the plant or the host organism. Methods for introducing nucleotide constructs into plants and other host cells are known in the art including, but not limited to, stable transformation methods, transient transformation methods, and virus-mediated methods.

The methods result in a transformed organisms, such as a plant, including whole plants, as well as plant organs (e.g., leaves, stems, roots, etc.), seeds, plant cells, propagules, embryos and progeny of the same. Plant cells can be differentiated or undifferentiated (e.g. callus, suspension culture cells, protoplasts, leaf cells, root cells, phloem cells, pollen).

“Transgenic plants” or “transformed plants” or “stably transformed” plants or cells or tissues refers to plants that have incorporated or integrated a polynucleotide encoding at least one pesticidal polypeptide of the invention. It is recognized that other exogenous or endogenous nucleic acid sequences or DNA fragments may also be incorporated into the plant cell. Agrobacterium- and biolistic-mediated transformation remain the two predominantly employed approaches. However, transformation may be performed by infection, transfection, microinjection, electroporation, microprojection, biolistics or particle bombardment, electroporation, silica/carbon fibers, ultrasound mediated, PEG mediated, calcium phosphate co-precipitation, polycation DMSO technique, DEAE dextran procedure, Agro and viral mediated (Caulimoriviruses, Geminiviruses, RNA plant viruses), liposome mediated and the like.

Transformation protocols as well as protocols for introducing polypeptides or polynucleotide sequences into plants may vary depending on the type of plant or plant cell, i.e., monocot or dicot, targeted for transformation. Methods for transformation are known in the art and include those set forth in U.S. Pat. Nos. 8,575,425; 7,692,068; 8,802,934; 7,541,517; each of which is herein incorporated by reference. See, also, Rakoczy-Trojanowska, M. (2002) Cell Mol Biol Lett. 7:849-858; Jones et al. (2005) Plant Methods 1:5; Rivera et al. (2012) Physics of Life Reviews 9:308-345; Bartlett et al. (2008) Plant Methods 4:1-12; Bates, G. W. (1999) Methods in Molecular Biology 111:359-366; Binns and Thomashow (1988) Annual Reviews in Microbiology 42:575-606; Christou, P. (1992) The Plant Journal 2:275-281; Christou, P. (1995) Euphytica 85:13-27; Tzfira et al. (2004) TRENDS in Genetics 20:375-383; Yao et al. (2006) Journal of Experimental Botany 57:3737-3746; Zupan and Zambryski (1995) Plant Physiology 107:1041-1047; Jones et al. (2005) Plant Methods 1:5.

Transformation may result in stable or transient incorporation of the nucleic acid into the cell. “Stable transformation” is intended to mean that the nucleotide construct introduced into a host cell integrates into the genome of the host cell and is capable of being inherited by the progeny thereof “Transient transformation” is intended to mean that a polynucleotide is introduced into the host cell and does not integrate into the genome of the host cell.

Methods for transformation of chloroplasts are known in the art. See, for example, Svab et al. (1990) Proc. Nail. Acad. Sci. USA 87:8526-8530; Svab and Maliga (1993) Proc. Natl. Acad. Sci. USA 90:913-917; Svab and Maliga (1993) EMBO J. 12:601-606. The method relies on particle gun delivery of DNA containing a selectable marker and targeting of the DNA to the plastid genome through homologous recombination. Additionally, plastid transformation can be accomplished by transactivation of a silent plastid-borne transgene by tissue-preferred expression of a nuclear-encoded and plastid-directed RNA polymerase. Such a system has been reported in McBride et al. (1994) Proc. Natl. Acad. Sci. USA 91:7301-7305.

The cells that have been transformed may be grown into plants in accordance with conventional ways. See, for example, McCormick et al. (1986) Plant Cell Reports 5:81-84. These plants may then be grown, and either pollinated with the same transformed strain or different strains, and the resulting hybrid having constitutive expression of the desired phenotypic characteristic identified. Two or more generations may be grown to ensure that expression of the desired phenotypic characteristic is stably maintained and inherited and then seeds harvested to ensure expression of the desired phenotypic characteristic has been achieved. In this manner, the present invention provides transformed seed (also referred to as “transgenic seed”) having a nucleotide construct of the invention, for example, an expression cassette of the invention, stably incorporated into their genome.

In specific embodiments, the sequences provide herein can be targeted to specific sites within the genome of the host cell or plant cell. Such methods include, but are not limited to, meganucleases designed against the plant genomic sequence of interest (D'Halluin et al. 2013 Plant Biotechnol J); CRISPR-Cas9, TALENs, and other technologies for precise editing of genomes (Feng, et al. Cell Research 23:1229-1232, 2013, Podevin, et al. Trends Biotechnology, online publication, 2013, Wei et al., J. Gen Genomics, 2013, Zhang et al (2013) WO 2013/026740); Cre-lox site-specific recombination (Dale et al. (1995) Plant J 7:649-659; Lyznik, et al. (2007) Transgenic Plant J 1:1-9; FLP-FRT recombination (Li et al. (2009) Plant Physiol 151:1087-1095); Bxb1-mediated integration (Yau et al. Plant J (2011) 701:147-166); zinc-finger mediated integration (Wright et al. (2005) Plant J 44:693-705); Cai et al. (2009) Plant Mol Biol 69:699-709); and homologous recombination (Lieberman-Lazarovich and Levy (2011) Methods Mol Biol 701: 51-65); Puchta (2002) Plant Mol Biol 48:173-182).

The sequence provided herein may be used for transformation of any plant species, including, but not limited to, monocots and dicots. Examples of plants of interest include, but are not limited to, corn (maize), sorghum, wheat, sunflower, tomato, crucifers, peppers, potato, cotton, rice, soybean, sugarbeet, sugarcane, tobacco, barley, and oilseed rape, Brassica sp., alfalfa, rye, millet, safflower, peanuts, sweet potato, cassaya, coffee, coconut, pineapple, citrus trees, cocoa, tea, banana, avocado, fig, guava, mango, olive, papaya, cashew, macadamia, almond, oats, vegetables, ornamentals, and conifers.

Vegetables include, but are not limited to, tomatoes, lettuce, green beans, lima beans, peas, and members of the genus Curcumis such as cucumber, cantaloupe, and musk melon. Ornamentals include, but are not limited to, azalea, hydrangea, hibiscus, roses, tulips, daffodils, petunias, carnation, poinsettia, and chrysanthemum. Preferably, plants of the present invention are crop plants (for example, maize, sorghum, wheat, sunflower, tomato, crucifers, peppers, potato, cotton, rice, soybean, sugarbeet, sugarcane, tobacco, barley, oilseed rape, etc.).

As used herein, the term plant includes plant cells, plant protoplasts, plant cell tissue cultures from which plants can be regenerated, plant calli, plant clumps, and plant cells that are intact in plants or parts of plants such as embryos, pollen, ovules, seeds, leaves, flowers, branches, fruit, kernels, ears, cobs, husks, stalks, roots, root tips, anthers, and the like. Grain is intended to mean the mature seed produced by commercial growers for purposes other than growing or reproducing the species. Progeny, variants, and mutants of the regenerated plants are also included within the scope of the invention, provided that these parts comprise the introduced polynucleotides. Further provided is a processed plant product or byproduct that retains the sequences disclosed herein, including for example, soymeal.

In another embodiment, the genes encoding the pesticidal proteins can be used to transform insect pathogenic organisms. Such organisms include baculoviruses, fungi, protozoa, bacteria, and nematodes. Microorganism hosts that are known to occupy the “phytosphere” (phylloplane, phyllosphere, rhizosphere, and/or rhizoplana) of one or more crops of interest may be selected. These microorganisms are selected so as to be capable of successfully competing in the particular environment with the wild-type microorganisms, provide for stable maintenance and expression of the gene expressing the pesticidal protein, and desirably, provide for improved protection of the pesticide from environmental degradation and inactivation.

Such microorganisms include archaea, bacteria, algae, and fungi. Of particular interest are microorganisms such as bacteria, e.g., Bacillus, Pseudomonas, Erwinia, Serratia, Klebsiella, Xanthomonas, Streptomyces, Rhizobium, Rhodopseudomonas, Methylius, Agrobacterium, Acetobacter, Lactobacillus, Arthrobacter, Azotobacter, Leuconostoc, and Alcaligenes. Fungi include yeast, e.g., Saccharomyces, Cryptococcus, Kluyveromyces, Sporobolomyces, Rhodotorula, and Aureobasidium. Of particular interest are such phytosphere bacterial species as Pseudomonas syringae, Pseudomonas aeruginosa, Pseudomonas fluorescens, Serratia marcescens, Acetobacter xylinum, Agrobacteria, Rhodopseudomonas spheroides, Xanthomonas campestris, Rhizobium melioti, Alcaligenes entrophus, Clavibacter xyli and Azotobacter vinlandir and phytosphere yeast species such as Rhodotorula rubra, R. glutinis, R. marina, R. aurantiaca, Cryptococcus albidus, C. diffluens, C. laurentii, Saccharomyces rosei, S. pretoriensis, S. cerevisiae, Sporobolomyces rosues, S. odorus, Kluyveromyces veronae, Aureobasidium pollulans, Bacillus thuringiensis, Escherichia coli, Bacillus subtilis, and the like.

Illustrative prokaryotes, both Gram-negative and gram-positive, include Enterobacteriaceae, such as Escherichia, Erwinia, Shigella, Salmonella, and Proteus; Bacillaceae; Rhizobiceae, such as Rhizobium; Spirillaceae, such as photobacterium, Zymomonas, Serratia, Aeromonas, Vibrio, Desulfovibrio, Spirillum; Lactobacillaceae; Pseudomonadaceae, such as Pseudomonas and Acetobacter; Azotobacteraceae and Nitrobacteraceae. Fungi include Phycomycetes and Ascomycetes, e.g., yeast, such as Saccharomyces and Schizosaccharomyces; and Basidiomycetes yeast, such as Rhodotorula, Aureobasidium, Sporobolomyces, and the like.

Genes encoding pesticidal proteins can be introduced by means of electrotransformation, PEG induced transformation, heat shock, transduction, conjugation, and the like. Specifically, genes encoding the pesticidal proteins can be cloned into a shuttle vector, for example, pHT3101 (Lerecius et al. (1989) FEMS Microbiol. Letts. 60: 211-218. The shuttle vector pHT3101 containing the coding sequence for the particular pesticidal protein gene can, for example, be transformed into the root-colonizing Bacillus by means of electroporation (Lerecius et al. (1989) FEMS Microbiol. Letts. 60: 211-218).

Expression systems can be designed so that pesticidal proteins are secreted outside the cytoplasm of gram-negative bacteria by fusing an appropriate signal peptide to the amino-terminal end of the pesticidal protein. Signal peptides recognized by E. coli include the OmpA protein (Ghrayeb et al. (1984) EMBO J, 3: 2437-2442).

Pesticidal proteins and active variants thereof can be fermented in a bacterial host and the resulting bacteria processed and used as a microbial spray in the same manner that Bacillus thuringiensis strains have been used as insecticidal sprays. In the case of a pesticidal protein(s) that is secreted from Bacillus, the secretion signal is removed or mutated using procedures known in the art. Such mutations and/or deletions prevent secretion of the pesticidal protein(s) into the growth medium during the fermentation process. The pesticidal proteins are retained within the cell, and the cells are then processed to yield the encapsulated pesticidal proteins.

Alternatively, the pesticidal proteins are produced by introducing heterologous genes into a cellular host. Expression of the heterologous gene results, directly or indirectly, in the intracellular production and maintenance of the pesticide. These cells are then treated under conditions that prolong the activity of the toxin produced in the cell when the cell is applied to the environment of target pest(s). The resulting product retains the toxicity of the toxin. These naturally encapsulated pesticidal proteins may then be formulated in accordance with conventional techniques for application to the environment hosting a target pest, e.g., soil, water, and foliage of plants. See, for example U.S. Pat. No. 6,468,523 and U.S. Publication No. 20050138685, and the references cited therein. In the present invention, a transformed microorganism (which includes whole organisms, cells, spore(s), pesticidal protein(s), pesticidal component(s), pest-impacting component(s), mutant(s), living or dead cells and cell components, including mixtures of living and dead cells and cell components, and including broken cells and cell components) or an isolated pesticidal protein can be formulated with an acceptable carrier into a pesticidal or agricultural composition(s) that is, for example, a suspension, a solution, an emulsion, a dusting powder, a dispersible granule, a wettable powder, and an emulsifiable concentrate, an aerosol, an impregnated granule, an adjuvant, a coatable paste, and also encapsulations in, for example, polymer substances.

Agricultural compositions may comprise a polypeptide, a recombinogenic polypeptide or a variant or fragment thereof, as disclosed herein. The agricultural composition disclosed herein may be applied to the environment of a plant or an area of cultivation, or applied to the plant, plant part, plant cell, or seed.

Such compositions disclosed above may be obtained by the addition of a surface-active agent, an inert carrier, a preservative, a humectant, a feeding stimulant, an attractant, an encapsulating agent, a binder, an emulsifier, a dye, a UV protectant, a buffer, a flow agent or fertilizers, micronutrient donors, or other preparations that influence plant growth. One or more agrochemicals including, but not limited to, herbicides, insecticides, fungicides, bactericides, nematicides, molluscicides, acaracides, plant growth regulators, harvest aids, and fertilizers, can be combined with carriers, surfactants or adjuvants customarily employed in the art of formulation or other components to facilitate product handling and application for particular target pests. Suitable carriers and adjuvants can be solid or liquid and correspond to the substances ordinarily employed in formulation technology, e.g., natural or regenerated mineral substances, solvents, dispersants, wetting agents, tackifiers, binders, or fertilizers. The active ingredients of the present invention are normally applied in the form of compositions and can be applied to the crop area, plant, or seed to be treated. For example, the compositions of the present invention may be applied to grain in preparation for or during storage in a grain bin or silo, etc. The compositions of the present invention may be applied simultaneously or in succession with other compounds. Methods of applying an active ingredient of the present invention or an agrochemical composition of the present invention that contains at least one of the pesticidal proteins produced by the bacterial strains of the present invention include, but are not limited to, foliar application, seed coating, and soil application. The number of applications and the rate of application depend on the intensity of infestation by the corresponding pest.

Suitable surface-active agents include, but are not limited to, anionic compounds such as a carboxylate of, for example, a metal; a carboxylate of a long chain fatty acid; an N-acylsarcosinate; mono or di-esters of phosphoric acid with fatty alcohol ethoxylates or salts of such esters; fatty alcohol sulfates such as sodium dodecyl sulfate, sodium octadecyl sulfate or sodium cetyl sulfate; ethoxylated fatty alcohol sulfates; ethoxylated alkylphenol sulfates; lignin sulfonates; petroleum sulfonates; alkyl aryl sulfonates such as alkyl-benzene sulfonates or lower alkylnaphtalene sulfonates, e.g., butyl-naphthalene sulfonate; salts of sulfonated naphthalene-formaldehyde condensates; salts of sulfonated phenol-formaldehyde condensates; more complex sulfonates such as the amide sulfonates, e.g., the sulfonated condensation product of oleic acid and N-methyl taurine; or the dialkyl sulfosuccinates, e.g., the sodium sulfonate of dioctyl succinate. Non-ionic agents include condensation products of fatty acid esters, fatty alcohols, fatty acid amides or fatty-alkyl- or alkenyl-substituted phenols with ethylene oxide, fatty esters of polyhydric alcohol ethers, e.g., sorbitan fatty acid esters, condensation products of such esters with ethylene oxide, e.g., polyoxyethylene sorbitar fatty acid esters, block copolymers of ethylene oxide and propylene oxide, acetylenic glycols such as 2,4,7,9-tetraethyl-5-decyn-4,7-diol, or ethoxylated acetylenic glycols. Examples of a cationic surface-active agent include, for instance, an aliphatic mono-, di-, or polyamine such as an acetate, naphthenate or oleate; or oxygen-containing amine such as an amine oxide of polyoxyethylene alkylamine; an amide-linked amine prepared by the condensation of a carboxylic acid with a di- or polyamine; or a quaternary ammonium salt.

Examples of inert materials include but are not limited to inorganic minerals such as kaolin, phyllosilicates, carbonates, sulfates, phosphates, or botanical materials such as cork, powdered corncobs, peanut hulls, rice hulls, and walnut shells.

The compositions of the present invention can be in a suitable form for direct application or as a concentrate of primary composition that requires dilution with a suitable quantity of water or other diluant before application. The pesticidal concentration will vary depending upon the nature of the particular formulation, specifically, whether it is a concentrate or to be used directly. The composition contains 1 to 98% of a solid or liquid inert carrier, and 0 to 50% or 0.1 to 50% of a surfactant. These compositions will be administered at the labeled rate for the commercial product, for example, about 0.01 lb-5.0 lb. per acre when in dry form and at about 0.01 pts.-10 pts. per acre when in liquid form.

In a further embodiment, the compositions, as well as the transformed microorganisms and pesticidal proteins, provided herein can be treated prior to formulation to prolong the pesticidal activity when applied to the environment of a target pest as long as the pretreatment is not deleterious to the pesticidal activity. Such treatment can be by chemical and/or physical means as long as the treatment does not deleteriously affect the properties of the composition(s). Examples of chemical reagents include but are not limited to halogenating agents; aldehydes such as formaldehyde and glutaraldehyde; anti-infectives, such as zephiran chloride; alcohols, such as isopropanol and ethanol; and histological fixatives, such as Bouin's fixative and Helly's fixative (see, for example, Humason (1967) Animal Tissue Techniques (W.H. Freeman and Co.).

In one aspect, pests may be killed or reduced in numbers in a given area by application of the pesticidal proteins of the invention to the area. Alternatively, the pesticidal proteins may be prophylactically applied to an environmental area to prevent infestation by a susceptible pest. Preferably the pest ingests, or is contacted with, a pesticidally-effective amount of the polypeptide. By “pesticidally-effective amount” is intended an amount of the pesticide that is able to bring about death to at least one pest, or to noticeably reduce pest growth, feeding, or normal physiological development. This amount will vary depending on such factors as, for example, the specific target pests to be controlled, the specific environment, location, plant, crop, or agricultural site to be treated, the environmental conditions, and the method, rate, concentration, stability, and quantity of application of the pesticidally-effective polypeptide composition. The formulations or compositions may also vary with respect to climatic conditions, environmental considerations, and/or frequency of application and/or severity of pest infestation.

The active ingredients are normally applied in the form of compositions and can be applied to the crop area, plant, or seed to be treated. Methods are therefore provided for providing to a plant, plant cell, seed, plant part or an area of cultivation, an effective amount of the agricultural composition comprising the polypeptide, recombinogenic polypeptide or an active variant or fragment thereof. By “effective amount” is intended an amount of a protein or composition sufficient to kill or control the pest or result in a noticeable reduction in pest growth, feeding, or normal physiological development. Such decreases in pest numbers, pest growth, pest feeding or pest normal development can comprise any statistically significant decrease, including, for example a decrease of about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 85%, 90%, 95% or greater. For example, the compositions may be applied to grain in preparation for or during storage in a grain bin or silo, etc. The compositions may be applied simultaneously or in succession with other compounds. Methods of applying an active ingredient or an agrochemical composition comprising at least one of the polypeptides, recombinogenic polypeptides or variants or fragments thereof as disclosed herein, include but are not limited to, foliar application, seed coating, and soil application.

Methods for increasing plant yield are provided. The methods comprise providing a plant or plant cell expressing a polynucleotide encoding the pesticidal polypeptide sequence disclosed herein and growing the plant or a seed thereof in a field infested with (or susceptible to infestation by) a pest against which said polypeptide has pesticidal activity. In some embodiments, the polypeptide has pesticidal activity against a lepidopteran, coleopteran, dipteran, hemipteran, or nematode pest, and said field is infested with a lepidopteran, hemipteran, coleopteran, dipteran, or nematode pest. As defined herein, the “yield” of the plant refers to the quality and/or quantity of biomass produced by the plant. By “biomass” is intended any measured plant product. An increase in biomass production is any improvement in the yield of the measured plant product. Increasing plant yield has several commercial applications. For example, increasing plant leaf biomass may increase the yield of leafy vegetables for human or animal consumption. Additionally, increasing leaf biomass can be used to increase production of plant-derived pharmaceutical or industrial products. An increase in yield can comprise any statistically significant increase including, but not limited to, at least a 1% increase, at least a 3% increase, at least a 5% increase, at least a 10% increase, at least a 20% increase, at least a 30%, at least a 50%, at least a 70%, at least a 100% or a greater increase in yield compared to a plant not expressing the pesticidal sequence. In specific methods, plant yield is increased as a result of improved pest resistance of a plant expressing a pesticidal protein disclosed herein. Expression of the pesticidal protein results in a reduced ability of a pest to infest or feed.

The plants can also be treated with one or more chemical compositions, including one or more herbicide, insecticides, or fungicides.

Non-limiting embodiments include:

1. An isolated polypeptide having insecticidal activity, comprising:

-   -   (a) a polypeptide comprising an amino acid sequence selected         from the group consisting of sequences set forth in SEQ ID         NOs:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,         18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,         34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49,         50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,         66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81,         82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,         98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,         111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123,         124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,         137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149,         150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162,         163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175,         176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188,         189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201,         202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214,         215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227,         228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240,         241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,         254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266,         267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279,         280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292,         293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,         306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318,         319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331,         332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,         345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357,         358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370,         371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,         384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396,         397, or 398; or     -   (b) a polypeptide comprising an amino acid sequence having at         least the percent sequence identity set forth in Table 1 to an         amino acid sequence selected from the group consisting of         sequences set forth in SEQ ID Nos: 1, 2, 3, 4, 5, 6, 7, 8, 9,         10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,         26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41,         42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57,         58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73,         74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89,         90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104,         105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117,         118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130,         131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143,         144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156,         157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169,         170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182,         183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195,         196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208,         209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221,         222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234,         235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247,         248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260,         261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273,         274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286,         287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299,         300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312,         313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325,         326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338,         339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351,         352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364,         365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377,         378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390,         391, 392, 393, 394, 395, 396, 397, or 398.

2. The polypeptide of embodiment 1, wherein said polypeptide comprises the amino acid sequence set forth in SEQ ID Nos. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398.

3. The polypeptide of embodiment 1 or 2, further comprising heterologous amino acid sequences.

4. A composition comprising the polypeptide of embodiments 1, 2, or 3.

5. A recombinant nucleic acid molecule that encodes the polypeptide of any one of embodiments 1 to 3, wherein said recombinant nucleic acid molecule is not the naturally occurring sequence encoding said polypeptide.

6. The recombinant nucleic acid of embodiment 5, wherein said nucleic acid molecule is a synthetic sequence that has been designed for expression in a plant.

7. The recombinant nucleic acid molecule of embodiment 5 or 6, wherein said nucleic acid molecule is operably linked to a promoter capable of directing expression in a plant cell.

8. The recombinant nucleic acid molecule of any one of embodiments 5 to 7, wherein said nucleic acid molecule is operably linked to a promoter capable of directing expression in a bacteria.

9. A host cell that contains the recombinant nucleic acid molecule of any one of embodiments 5 to 8.

10. The host cell of embodiment 9, wherein said host cell is a bacterial host cell.

11. A DNA construct comprising a promoter that drives expression in a plant cell operably linked to a recombinant nucleic acid molecule comprising:

-   -   (a) a nucleotide sequence that encodes a polypeptide comprising         the amino acid sequence of any one of SEQ ID NOs:1, 2, 3, 4, 5,         6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,         23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38,         39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54,         55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,         71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86,         87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101,         102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114,         115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127,         128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140,         141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153,         154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166,         167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179,         180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,         193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205,         206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218,         219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231,         232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244,         245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257,         258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270,         271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283,         284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296,         297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309,         310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322,         323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335,         336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348,         349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361,         362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374,         375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387,         388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398; or,     -   (b) a nucleotide sequence that encodes a polypeptide comprising         an amino acid sequence having at least the percent sequence         identity set forth in Table 1 to an amino acid sequence selected         from the group consisting of sequences set forth in SEQ ID         NOs:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,         18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,         34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49,         50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,         66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81,         82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,         98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,         111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123,         124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,         137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149,         150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162,         163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175,         176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188,         189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201,         202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214,         215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227,         228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240,         241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,         254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266,         267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279,         280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292,         293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,         306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318,         319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331,         332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,         345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357,         358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370,         371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,         384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396,         397, or 398.

12. The DNA construct of embodiment 11, wherein said nucleotide sequence is a synthetic DNA sequence that has been designed for expression in a plant.

13. A vector comprising the DNA construct of embodiment 11 or 12.

14. A host cell that contains the DNA construct of any one of embodiments 11-13 or the vector of embodiment 13.

15. The host cell of embodiment 14, wherein the host cell is a plant cell.

16. A transgenic plant comprising the host cell of embodiment 15.

17. A composition comprising the host cell of any one of embodiments 9, 10, 14, or 15.

18. The composition of embodiment 17, wherein said composition is selected from the group consisting of a powder, dust, pellet, granule, spray, emulsion, colloid, and solution.

19. The composition of embodiment 17 or 18, wherein said composition comprises from about 1% to about 99% by weight of said polypeptide.

20. A method for controlling a pest population comprising contacting said population with a pesticidal-effective amount of the composition of any one of embodiments 4 or 17-19.

21. A method for killing a pest population comprising contacting said population with a pesticidal-effective amount of the composition of any one of embodiments 4 or 17-19.

22. A method for producing a polypeptide with pesticidal activity, comprising culturing the host cell of any one of embodiments 9, 10, 14, or 15 under conditions in which the nucleic acid molecule encoding the polypeptide is expressed.

23. A plant having stably incorporated into its genome a DNA construct comprising a nucleotide sequence that encodes a protein having pesticidal activity, wherein said nucleotide sequence comprise:

-   -   (a) a nucleotide sequence that encodes a polypeptide comprising         the amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5,         6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,         23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38,         39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54,         55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,         71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86,         87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101,         102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114,         115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127,         128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140,         141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153,         154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166,         167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179,         180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,         193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205,         206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218,         219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231,         232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244,         245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257,         258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270,         271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283,         284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296,         297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309,         310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322,         323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335,         336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348,         349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361,         362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374,         375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387,         388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398; or,     -   (b) a nucleotide sequence that encodes a polypeptide comprising         an amino acid sequence having at least the percent sequence         identity set forth in Table 1 to an amino acid sequence selected         from the group consisting of sequences set forth in SEQ ID         NOs:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,         18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,         34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49,         50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,         66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81,         82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,         98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,         111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123,         124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,         137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149,         150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162,         163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175,         176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188,         189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201,         202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214,         215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227,         228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240,         241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,         254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266,         267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279,         280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292,         293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,         306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318,         319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331,         332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,         345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357,         358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370,         371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,         384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396,         397, or 398.

24. A transgenic seed of the plant of embodiment 23.

25. A method for protecting a plant from an insect pest, comprising expressing in a plant or cell thereof a nucleotide sequence that encodes a pesticidal polypeptide, wherein said nucleotide sequence comprising:

-   -   (a) a nucleotide sequence that encodes a polypeptide comprising         the amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5,         6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,         23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38,         39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54,         55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,         71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86,         87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101,         102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114,         115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127,         128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140,         141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153,         154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166,         167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179,         180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,         193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205,         206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218,         219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231,         232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244,         245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257,         258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270,         271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283,         284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296,         297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309,         310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322,         323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335,         336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348,         349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361,         362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374,         375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387,         388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398; or,     -   (b) a nucleotide sequence that encodes a polypeptide comprising         an amino acid sequence having at least the percent sequence         identity set forth in Table 1 to an amino acid sequence selected         from the group consisting of sequences set forth in SEQ ID         NOs:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,         18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,         34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49,         50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,         66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81,         82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,         98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,         111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123,         124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,         137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149,         150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162,         163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175,         176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188,         189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201,         202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214,         215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227,         228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240,         241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,         254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266,         267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279,         280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292,         293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,         306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318,         319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331,         332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,         345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357,         358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370,         371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,         384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396,         397, or 398.

26. The method of embodiment 25, wherein said plant produces a pesticidal polypeptide having pesticidal activity against a lepidopteran or coleopteran pest.

27. A method for increasing yield in a plant comprising growing in a field a plant or seed thereof having stably incorporated into its genome a DNA construct comprising a promoter that drives expression in a plant operably linked to a nucleotide sequence that encodes a pesticidal polypeptide, wherein said nucleotide sequence comprises:

-   -   (a) a nucleotide sequence that encodes a polypeptide comprising         the amino acid sequence of any one of SEQ ID NOs:1, 2, 3, 4, 5,         6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,         23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38,         39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54,         55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,         71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86,         87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101,         102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114,         115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127,         128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140,         141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153,         154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166,         167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179,         180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,         193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205,         206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218,         219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231,         232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244,         245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257,         258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270,         271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283,         284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296,         297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309,         310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322,         323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335,         336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348,         349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361,         362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374,         375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387,         388, 389, 390, 391, 392, 393, 394, 395, 396, 397, or 398; or,     -   (b) a nucleotide sequence that encodes a polypeptide comprising         an amino acid sequence having at least the percent sequence         identity set forth in Table 1 to an amino acid sequence selected         from the group consisting of sequences set forth in SEQ ID         NOs:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,         18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33,         34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49,         50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,         66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81,         82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,         98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,         111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123,         124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,         137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149,         150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162,         163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175,         176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188,         189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201,         202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214,         215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227,         228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240,         241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,         254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266,         267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279,         280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292,         293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,         306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318,         319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331,         332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,         345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357,         358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370,         371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,         384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396,         397, or 398.

28. A method of obtaining a polynucleotide that encodes an improved polypeptide comprising pesticidal activity is provided, wherein the improved polypeptide has at least one improved property over any one of SEQ ID NOS: 1-398 comprising:

-   -   (a) recombining a plurality of parental polynucleotides         comprising SEQ ID NO: 1-398 or an active variant or fragment         thereof to produce a library of recombinant polynucleotides         encoding recombinant pesticidal polypeptides;     -   (b) screening the library to identify a recombinant         polynucleotide that encodes an improved recombinant pesticidal         polypeptide that has an enhanced property improved over the         parental polynucleotide;     -   (c) recovering the recombinant polynucleotide that encodes the         improved recombinant pesticidal polypeptide identified in (b);         and,     -   (d) repeating steps (a), (b) and (c) using the recombinant         polynucleotide recovered in step (c) as one of the plurality of         parental polynucleotides in repeated step (a).

The following examples are offered by way of illustration and not by way of limitation.

EXPERIMENTAL Experiment 1: Discovery of Novel Genes by Sequencing and DNA Analysis

Microbial cultures were grown in liquid culture in standard laboratory media. Cultures were grown to saturation (16 to 24 hours) before DNA preparation. DNA was extracted from bacterial cells by detergent lysis, followed by binding to a silica matrix and washing with an ethanol buffer. Purified DNA was eluted from the silica matrix with a mildly alkaline aqueous buffer.

DNA for sequencing was tested for purity and concentration by spectrophotometry. Sequencing libraries were prepared using the Nextera XT library preparation kit according to the manufacturer's protocol. Sequence data was generated on a HiSeq 2000 according to the Illumina HiSeq 2000 System User Guide protocol.

Sequencing reads were assembled into draft genomes using the CLC Bio Assembly Cell software package. Following assembly, gene calls were made by several methods and resulting gene sequences were interrogated to identify novel homologs of pesticidal genes. Novel genes were identified by BLAST, by domain composition, and by pairwise alignment versus a target set of pesticidal genes. A summary of such sequences is set forth in Table 1.

Genes identified in the homology search were amplified from bacterial DNA by PCR and cloned into bacterial expression vectors containing fused in-frame purification tags. Cloned genes were expressed in E. coli and purified by column chromatography. Purified proteins were assessed in insect diet bioassay studies to identify active proteins.

Insect diet bioassays were performed using a wheat germ and agar artificial diet to which purified protein were applied as a surface treatment. Insect larvae were applied to treated diet and monitored for mortality.

Insect diet bioassays were performed using a sucrose liquid diet contained in a membrane sachet to which purified protein was added. Insect nymphs were allowed to feed on the diet sachet and were monitored for mortality. Insects tested in bioassays included the Brown Stink Bug (BSB), Euschistus servus, and the Southern Green Stink Bug (SGSB), Nezara viridula.

Example 2. Heterologous Expression in E. coli

Each open reading frame set forth in Tables 3 and 4 was cloned into an E. coli expression vector containing a maltose binding protein (pMBP). The expression vector was transformed into BL21*RIPL. An LB culture supplemented with carbenicillin was inoculated with a single colony and grown overnight at 37° C. using 0.5% of the overnight culture, a fresh culture was inoculated and grown to logarithmic phase at 37° C. The culture was induced using 250 mM IPTG for 18 hours at 16° C. The cells were pelleted and resuspended in 10 mM Tris pH7.4 and 150 mM NaCl supplemented with protease inhibitors. The protein expression was evaluated by SDS-PAGE.

Example 3. Pesticidal Activity Against Coleopteran and Lepidoptera

Protein Expression: Each sequence set forth in Table 3 was expressed in E. coli as described in Example 2. 400 mL of LB was inoculated and grown to an OD600 of 0.6. The culture was induced with 0.25 mM IPTG overnight at 16° C. The cells were spun down and the cell pellet was resuspend in 5 mL of buffer. The resuspension was sonicated for 2 min on ice.

Bioassay: Fall army worm (FAW), corn ear worm (CEW), European corn borer (ECB) southwestern corn borer (SWCB) and diamond backed moth (DBM or Px) eggs were purchased from a commercial insectary (Benzon Research Inc., Carlisle, Pa.). The FAW, CEW, ECB and BCW eggs were incubated to the point that eclosion would occur within 12 hrs of the assay setup. SWCB and DBM were introduced to the assay as neonate larvae. Assays were carried out in 24-well trays containing multispecies lepidopteran diet (Southland Products Inc., Lake Village, Ark.). Samples of the sonicated lysate were applied to the surface of the diet (diet overlay) and allowed to evaporate and soak into the diet. For CEW, FAW, BCW, ECB and SWCB, a 125 μl of sonicated lysate was added to the diet surface and dried. For DBM, 50 μl of a 1:2 dilution of sonicated lysate was added to the diet surface. The bioassay plates were sealed with a plate sealing film vented with pin holes. The plates were incubated at 26° C. at 65% relative humidity (RH) on a 16:8 day:night cycle in a Percival for 5 days. The assays were assessed for level of mortality, growth inhibition and feeding inhibition.

For the western corn rootworm bioassay, the protein construct/lysate was evaluated in an insect bioassay by dispensing 60 μl volume on the top surface of diet in well/s of 24-well plate (Cellstar, 24-well, Greiner Bio One) and allowed to dry. Each well contained 500 μl diet (Marrone et al., 1985). Fifteen to twenty neonate larvae were introduced in each well using a fine tip paint brush and the plate was covered with membrane (Viewseal, Greiner Bio One). The bioassay was stored at ambient temperature and scored for mortality, and/or growth/feeding inhibition at day 4.

For Colorado Potato Beetle (CPB) a cork bore size No. 8 leaf disk was excised from potato leaf and was dipped in the protein construct/lysate until thoroughly wet and placed on top of filter disk (Millipore, glass fiber filter, 13 mm). 60 μl dH2O was added to each filter disk and placed in each well of 24-well plate (Cellstar, 24-well, Greiner Bio One). The leaf disk was allowed to dry and five to seven first instar larvae were introduced in each well using a fine tip paint brush. The plate was covered with membrane (Viewseal, Greiner Bio One) and small hole was punctured in each well of the membrane. The construct was evaluated with four replicates, and scored for mortality and leaf damage on day 3.

Table 3 provides a summary of pesticidal activity against coleopteran and lepidoptera of the various sequences. Table code: “−” indicates no activity seen; “+” indicates pesticidal activity; “NT” indicates not tested; “S” indicates stunt; “SS” indicates slight stunt; “LF” indicates low feeding, “M” indicates mortality.

TABLE 3 Summary of Pesticidal Activity against Coleopteran and Lepidoptera. WCR Mortality APG Seq ID FAW CEW BCW ECB SWCB CPB Px (%) APG00056 1 − − − − − − − 80-100% mortality APG00073 11 S − − − − NT − 80-100% mortality APG00105 18 − − − − − NT − NT APG00121 29 − − − − − NT − 0-50% mortality APG00131 30 − − − − − NT − NT APG00152 36 M, S − − − − − NT 50-80% mortality APG00164 41 − − − − − + NT 0-50% mortality APG00175 49 SS − − − − NT + NT APG00176 50 − − − − − + + 80-100% mortality APG00342 142 − − − − − − − 80-100% mortality APG00388 159 NT NT NT NT NT − + NT APG00799 159 SS SS − − − NT − NT APG00801 160 − − − − − − − 0-50% mortality APG00422 177 − − − − M, SS + + NT APG00456 180 − − − − − NT − 0-50% mortality APG00462 183 NT NT NT NT NT + + NT APG00084 229 − SS − NT NT NT NT 0-50% mortality APG00108 230 − − − − − NT − 60-100% mortality APG00116 233 HM, S − − − SS NT − 80-100% mortality APG00134 240 − − − − − NT − 60-100% mortality APG00168 247 NT NT NT NT NT − + NT APG00177 248 M, S NT NT NT S NT − 60-100% mortality APG00186 249 − − − NT − 0-50% mortality APG00194 251 SS − − − − NT − 80-100% mortality APG00200 254 NT NT NT NT NT + + NT APG00210 258 M, S − − − − NT NT 80-100% mortality APG00227 267 − − − − − NT NT 0-50% mortality APG00239 277 M, S − − − − NT NT 80-100% mortality APG00248 282 NT NT NT NT NT + NT NT APG00267 293 − − − − − − NT 50-80% mortality APG00273 296 − − − − SS NT − NT APG00290 307 M, S − − − HM, S NT NT 80-100% mortality APG00291 308 M, S − − − − NT NT 80-100% mortality APG00297 313 M, S − − − − NT NT 50-80% mortality APG00500 357 − − − − − − + 60-100% mortality APG00647 372 − − − − M, SS NT − NT APG00698 374 − − − − M, SS NT + NT

Example 4. Pesticidal Activity Against Hemipteran

Protein Expression: Each of the sequences set forth in Table 4 was expressed in E. coli as described in Example 2. 400 mL of LB was inoculated and grown to an OD600 of 0.6. The culture was induced with 0.25 mM IPTG overnight at 16° C. The cells were spun down and the cell pellet was re-suspend in 5 mL of buffer. The resuspension was sonicated for 2 min on ice.

Second instar SGSB were obtained from a commercial insectary (Benzon Research Inc., Carlisle, Pa.). A 50% v/v ratio of sonicated lysate sample to 20% sucrose was employed in the bioassay. Stretched parafilm was used as a feeding membrane to expose the SGSB to the diet/sample mixture. The plates were incubated at 25° C.:21° C., 16:8 day:night cycle at 65% RH for 5 days.

Mortality was scored for each sample. The results are set forth in Table 4. A dashed line indicates no mortality was detected. The proteins listed in table 4 showed 25% mortality or 50% mortality (as indicated) against southern green stinkbug (1 stinkbug out of 4 died). The negative controls (empty vector expressed binding domain and buffer only) both showed no mortality (0 stinkbugs out of 4).

TABLE 4 Summary of Pesticidal Activity against Hemipteran APG Seq ID Tested against SGSB APG00173 47 25% APG00190 62 25% APG00388 159 25% APG00801 160 50% APG00196 252 50% APG00273 296 50% APG00291 308 25% APG00297 313 25%

Example 5. Transformation of Soybean

DNA constructs comprising each of SEQ ID NOs: 1-398 or active variants or fragments thereof operably linked to a promoter active in a plant are cloned into transformation vectors and introduced into Agrobacterium as described in PCT Application PCT/US2015/066702, filed Dec. 18, 2015, herein incorporated by reference in its entirety.

Four days prior to inoculation, several loops of Agrobacterium are streaked to a fresh plate of YEP* medium supplemented with the appropriate antibiotics** (spectinomycin, chloramphenicol and kanamycin). Bacteria are grown for two days in the dark at 28° C. After two days, several loops of bacteria are transferred to 3 ml of YEP liquid medium with antibiotics in a 125 ml Erlenmeyer flask. Flasks are placed on a rotary shaker at 250 RPM at 28° C. overnight. One day before inoculation, 2-3 ml of the overnight culture were transferred to 125 ml of YEP with antibiotics in a 500 ml Erlenmeyer flask. Flasks are placed on a rotary shaker at 250 RPM at 28° C. overnight.

Prior to inoculation, the OD of the bacterial culture is checked at OD 620. An OD of 0.8-1.0 indicates that the culture is in log phase. The culture is centrifuged at 4000 RPM for 10 minutes in Oakridge tubes. The supernatant is discarded and the pellet is resuspended in a volume of Soybean Infection Medium (SI) to achieve the desired OD. The cultures are held with periodic mixing until needed for inoculation.

Two or three days prior to inoculation, soybean seeds are surface sterilized using chlorine gas. In a fume hood, a petri dish with seeds is placed in a bell jar with the lid off. 1.75 ml of 12 N HCl is slowly added to 100 ml of bleach in a 250 ml Erlenmeyer flask inside the bell jar. The lid is immediately placed on top of the bell jar. Seeds are allowed to sterilize for 14-16 hours (overnight). The top is removed from the bell jar and the lid of the petri dish is replaced. The petri dish with the surface sterilized is then opened in a laminar flow for around 30 minutes to disperse any remaining chlorine gas.

Seeds are imbibed with either sterile DI water or soybean infection medium (SI) for 1-2 days. Twenty to 30 seeds are covered with liquid in a 100×25 mm petri dish and incubated in the dark at 24° C. After imbibition, non-germinating seeds are discarded.

Cotyledonary explants are processed on a sterile paper plate with sterile filter paper dampened using SI medium employing the methods of U.S. Pat. No. 7,473,822, herein incorporated by reference.

Typically, 16-20 cotyledons are inoculated per treatment. The SI medium used for holding the explants is discarded and replaced with 25 ml of Agrobacterium culture (OD 620=0.8-20). After all explants are submerged, the inoculation is carried out for 30 minutes with periodic swirling of the dish. After 30 minutes, the Agrobacterium culture is removed.

Co-cultivation plates are prepared by overlaying one piece of sterile paper onto Soybean Co-cultivation Medium (SCC). Without blotting, the inoculated cotyledons are cultured adaxial side down on the filter paper. Around 20 explants can be cultured on each plate. The plates are sealed with Parafilm and cultured at 24° C. and around 120 μmoles m-2s-1 (in a Percival incubator) for 4-5 days.

After co-cultivation, the cotyledons are washed 3 times in 25 ml of Soybean Wash Medium with 200 mg/l of cefotaxime and timentin. The cotyledons are blotted on sterile filter paper and then transferred to Soybean Shoot Induction Medium (SSI). The nodal end of the explant is depressed slightly into the medium with distal end kept above the surface at about 45 deg. No more than 10 explants are cultured on each plate. The plates are wrapped with Micropore tape and cultured in the Percival at 24° C. and around 120 μmoles m-2s-1.

The explants are transferred to fresh SSI medium after 14 days. Emerging shoots from the shoot apex and cotyledonary node are discarded. Shoot induction is continued for another 14 days under the same conditions.

After 4 weeks of shoot induction, the cotyledon is separated from the nodal end and a parallel cut is made underneath the area of shoot induction (shoot pad). The area of the parallel cut is placed on Soybean Shoot Elongation Medium (SSE) and the explants cultured in the Percival at 24° C. and around 120 μmoles m-2s-1. This step is repeated every two weeks for up to 8 weeks as long as shoots continue to elongate.

When shoots reach a length of 2-3 cm, they are transferred to Soybean Rooting Medium (SR) in a Plantcon vessel and incubated under the same conditions for 2 weeks or until roots reach a length of around 3-4 cm. After this, plants are transferred to soil.

Note, all media mentioned for soybean transformation are found in Paz et al. (2010) Agrobacterium-mediated transformation of soybean and recovery of transgenic soybean plants; Plant Transformation Facility of Iowa State University, which is herein incorporated by reference in its entirety.

Example 6. Transformation of Maize

Maize ears are best collected 8-12 days after pollination. Embryos are isolated from the ears, and those embryos 0.8-1.5 mm in size are preferred for use in transformation. Embryos are plated scutellum side-up on a suitable incubation media, such as DN62A5S media (3.98 g/L N6 Salts; 1 mL/L (of 1000× Stock) N6 Vitamins; 800 mg/L L-Asparagine; 100 mg/L Myo-inositol; 1.4 g/L L-Proline; 100 mg/L Casamino acids; 50 g/L sucrose; 1 mL/L (of 1 mg/mL Stock) 2,4-D). However, media and salts other than DN62A5S are suitable and are known in the art. Embryos are incubated overnight at 25° C. in the dark. However, it is not necessary per se to incubate the embryos overnight.

The resulting explants are transferred to mesh squares (30-40 per plate), transferred onto osmotic media for about 30-45 minutes, and then transferred to a beaming plate (see, for example, PCT Publication No. WO/0138514 and U.S. Pat. No. 5,240,842). DNA constructs designed to express the GRG proteins of the present invention in plant cells are accelerated into plant tissue using an aerosol beam accelerator, using conditions essentially as described in PCT Publication No. WO/0138514. After beaming, embryos are incubated for about 30 min on osmotic media, and placed onto incubation media overnight at 25° C. in the dark. To avoid unduly damaging beamed explants, they are incubated for at least 24 hours prior to transfer to recovery media. Embryos are then spread onto recovery period media, for about 5 days, 25° C. in the dark, and then transferred to a selection media. Explants are incubated in selection media for up to eight weeks, depending on the nature and characteristics of the particular selection utilized. After the selection period, the resulting callus is transferred to embryo maturation media, until the formation of mature somatic embryos is observed. The resulting mature somatic embryos are then placed under low light, and the process of regeneration is initiated by methods known in the art. The resulting shoots are allowed to root on rooting media, and the resulting plants are transferred to nursery pots and propagated as transgenic plants.

Example 7. Pesticidal Activity Against Nematodes

Heterodera Glycine's (Soybean Cyst Nematode) In Vitro Assay

Soybean Cyst Nematodes are dispensed into a 96 well assay plate with a total volume of 100 uls and 100 J2 per well. The protein of interest as set forth in any one of SEQ ID NOs: 1-398 is dispensed into the wells and held at room temperature for assessment. Finally, the 96 well plate containing the SCN J2 is analyzed for motility. Data is reported as % inhibition as compared to the controls. Hits are defined as greater or equal to 70% inhibition.

Heterodera Glycine's (Soybean Cyst Nematode) on Plant Assay

Soybean plants expressing one or more of SEQ ID NOs: 1-398 are generated as described elsewhere herein. A 3-week-old soybean cutting is inoculated with 5000 SCN eggs per plant. This infection is held for 70 days and then harvested for counting of SCN cyst that has developed on the plant. Data is reported as % inhibition as compared to the controls. Hits are defined as greater or equal to 90% inhibition.

Meloidogyne incognita (Root-Knot Nematode) In Vitro Assay

Root-Knot Nematodes are dispensed into a 96 well assay plate with a total volume of 100 uls and 100 J2 per well. The protein of interest comprising any one of SEQ ID NOs: 1-398 is dispensed into the wells and held at room temperature for assessment. Finally the 96 well plate containing the RKN J2 is analyzed for motility. Data is reported as % inhibition as compared to the controls. Hits are defined as greater or equal to 70% inhibition.

Meloidogyne incognita (Root-Knot Nematode) on Plant Assay

Soybean plants expressing one or more of SEQ ID NOs: 1-398 are generated as described elsewhere herein. A 3-week-old soybean is inoculated with 5000 RKN eggs per plant. This infection is held for 70 days and then harvested for counting of RKN eggs that have developed in the plant. Data is reported as % inhibition as compared to the controls. Hits are defined as greater or equal to 90% inhibition.

Example 8. Additional Assays for Pesticidal Activity

The various polypeptides set forth in SEQ ID NOs: 1-398 can be tested to act as a pesticide upon a pest in a number of ways. One such method is to perform a feeding assay. In such a feeding assay, one exposes the pest to a sample containing either compounds to be tested or control samples. Often this is performed by placing the material to be tested, or a suitable dilution of such material, onto a material that the pest will ingest, such as an artificial diet. The material to be tested may be composed of a liquid, solid, or slurry. The material to be tested may be placed upon the surface and then allowed to dry. Alternatively, the material to be tested may be mixed with a molten artificial diet, and then dispensed into the assay chamber. The assay chamber may be, for example, a cup, a dish, or a well of a microtiter plate.

Assays for sucking pests (for example aphids) may involve separating the test material from the insect by a partition, ideally a portion that can be pierced by the sucking mouth parts of the sucking insect, to allow ingestion of the test material. Often the test material is mixed with a feeding stimulant, such as sucrose, to promote ingestion of the test compound.

Other types of assays can include microinjection of the test material into the mouth, or gut of the pest, as well as development of transgenic plants, followed by test of the ability of the pest to feed upon the transgenic plant. Plant testing may involve isolation of the plant parts normally consumed, for example, small cages attached to a leaf, or isolation of entire plants in cages containing insects.

Other methods and approaches to assay pests are known in the art, and can be found, for example in Robertson and Preisler, eds. (1992) Pesticide bioassays with arthropods, CRC, Boca Raton, Fla. Alternatively, assays are commonly described in the journals Arthropod Management Tests and Journal of Economic Entomology or by discussion with members of the Entomological Society of America (ESA). Any one of SEQ ID NOS: 1-398 can be expressed and employed in an assay as set forth in Examples 3 and 4, herein.

All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this invention pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be obvious that certain changes and modifications may be practiced within the scope of the appended claims. 

That which is claimed is:
 1. A recombinant polypeptide having pesticidal activity, comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or 374, wherein the polypeptide has pesticidal activity; and wherein the polypeptide further comprises a heterologous amino acid chemically linked to the polypeptide.
 2. A composition comprising the polypeptide of claim
 1. 3. A recombinant nucleic acid molecule encoding a polypeptide comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or 374, wherein the polypeptide has pesticidal activity; and wherein said recombinant nucleic acid molecule is operably linked to a heterologous promoter.
 4. The recombinant nucleic acid of claim 3, wherein said nucleic acid molecule is a synthetic sequence designed for expression in a plant.
 5. The recombinant nucleic acid molecule of claim 3, wherein said nucleic acid molecule is operably linked to a promoter capable of directing expression in a plant cell.
 6. The recombinant nucleic acid molecule of claim 3, wherein said heterologous promoter is capable of directing expression in a bacterium.
 7. A host cell comprising the recombinant nucleic acid molecule of claim
 3. 8. The host cell of claim 7, wherein said host cell is a bacterial host cell.
 9. A DNA construct comprising a promoter that drives expression in a plant cell operably linked to a recombinant nucleic acid molecule comprising a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or
 374. 10. The DNA construct of claim 9, wherein said nucleotide sequence is a synthetic DNA sequence designed for expression in a plant.
 11. A vector comprising the DNA construct of claim
 9. 12. A host cell comprising the vector of claim
 11. 13. A composition comprising the host cell of claim
 12. 14. The composition of claim 13, wherein said composition is selected from the group consisting of a powder, dust, pellet, granule, spray, emulsion, colloid, and solution.
 15. The composition of claim 14, wherein said composition comprises from about 1% to about 99% by weight of said polypeptide.
 16. A method for controlling a pest population comprising contacting said pest population with a pesticidal-effective amount of the composition of claim
 2. 17. A method for producing a polypeptide with pesticidal activity comprising culturing the host cell of claim 12 under conditions in which the nucleic acid molecule encoding the polypeptide is expressed.
 18. A plant having stably incorporated into its genome a DNA construct comprising a nucleic acid molecule that encodes a protein having pesticidal activity, wherein said nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or 374, wherein the polypeptide has pesticidal activity.
 19. A transgenic seed of the plant of claim
 18. 20. A method for protecting a plant from an insect pest, comprising expressing in a plant or cell thereof a nucleic acid molecule that encodes a pesticidal polypeptide, wherein said nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or 374, wherein the polypeptide has pesticidal activity.
 21. The method of claim 20, wherein said plant produces a pesticidal polypeptide having pesticidal activity against at least one of a lepidopteran pest, a coleopteran pest, or a hemipteran pest.
 22. A method for increasing yield in a plant comprising growing in a field a plant or seed thereof having stably incorporated into its genome a DNA construct comprising a promoter that drives expression in a plant operably linked to a nucleic acid molecule that encodes a pesticidal polypeptide, wherein said nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence set forth in SEQ ID NOs: 206 or 374, wherein the polypeptide has pesticidal activity.
 23. The polypeptide of claim 1, wherein the polypeptide comprises the amino acid sequence set forth in SEQ ID NOs: 206 or
 374. 24. The recombinant nucleic acid molecule of claim 3, wherein the recombinant nucleic acid molecule encodes the amino acid sequence set forth in SEQ ID NOs: 206 or
 374. 25. The DNA construct of claim 9, wherein the nucleotide sequence encodes a polypeptide comprising the amino acid sequence set forth in SEQ ID NOs: 206 or
 374. 26. The plant of claim 18, wherein the nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising the amino acid sequence of any one of SEQ ID NOs: 206 or
 374. 27. The method of claim 20, wherein the nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising the amino acid sequence of any one of SEQ ID NOs: 206 or
 374. 28. The method of claim 22, wherein the nucleic acid molecule comprises a nucleotide sequence that encodes a polypeptide comprising the amino acid sequence of any one of SEQ ID NOs: 206 or
 374. 